Follow up of Learning Early About Peanut Allergy (LEAP) Participants and Their Families: LEAP Trio (ITN070AD)
Conditions: Peanut Allergy
Sponsor: National Institute of Allergy and Infectious Diseases (NIAID)

 
 
 
 
Immune  
Tolerance 
Network 
 
Follow up of LEAP Participants and Their Families: LEAP Trio 
 
Protocol ITN070AD 
Sponsored by: The National Institute of Allergy & Infectious 
Diseases (USA) 
Version 7.0 (10 December 2021) 
IND #: Not applicable 
Protocol Chair 
Gideon Lack, MD 
Professor of Paediatric Allergy 
Paediatric Allergy Clinical Trials Unit Level 1 
Evelina Children's Hospital Guy’s and St. Thomas’ 
NHS Foundation Trust 
Lambeth Palace Road London SE1 7EH 
Phone: (44) 0207 188 9730 Fax: (44) 0207 188 9782 
E-mail: gideon.lack@kcl.ac.uk 
ITN Clinical Trial Physician 
Michelle Huffaker, MD 
Clinical Trials Physician, Clinical Trials Group 
Immune Tolerance Network 
University of California, San Francisco 
Diabetes Center 
513 Parnassus Avenue, HSW 11, Box 0534 
San Francisco, CA 94143-0534 
Tel: 415-319-4303 
Email: mhuffaker@immunetolerance.org 
 
NIAID Medical Monitor 
Lisa M. Wheatley, MD, MPH 
Chief, Asthma and Airways Biology Section  
Division of Allergy, Immunology,  & 
Transplantation  
National Institute of Allergy & Infectious Diseases 
5601 Fishers Lane., Room 6B50 
Bethesda, MD, 20892-9827 
Tel: 240-627-3573 
Fax: 301-480-4258 
Email: lisa.wheatley@nih.gov 
 
ITN Biomarker & Discovery Research  
Carolyn H. Baloh, MD 
Biologist 
Allergy Assessment Group 
Immune Tolerance Network   
Brigham and Women’s Hospital 
60 Fenwood Rd 
BTM/Hale Building, 5th floor 
Boston, MA 02115 
Email: cbaloh@immunetolerance.org 
Rho Scientist 
Samuel Arbes, DDS, MPH, PhD  
Senior Research Scientist 
Rho  
2635 East NC Highway 54 
Durham, NC 27713  
Tel: 919-595-6570 
Fax: 919-408-0999 
Email: sam_arbes@rhoworld.com  

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LEAP Trio, IRAS ID 195451 
NIAID Senior Regulatory Officer 
Lyudmila Lyakh, MD, PhD 
Regulatory Officer 
Division of Allergy, Immunology, & 
Transplantation 
National Institute of Allergy & Infectious Diseases 
5601 Fishers Lane., Room 7B39 
Bethesda, MD, 20892-9828 
Tel: 240-627-3530 
Fax: 301-480-1537 
E-mail: lyudmill@niaid.nih.gov 
 
Rho Biostatistician 
Margie Byron, MStat 
Senior Biostatistician 
Rho  
2635 East NC Highway 54 
Durham, NC 27713  
Tel: 919.595.6454 
Email: margie_byron@rhoworld.com 
 
NIAID Project Manager 
Joy Laurienzo Panza, RN, BSN 
NIAID Project Manager 
Division of Allergy, Immunology, & 
Transplantation 
National Institute of Allergy & Infectious Diseases 
5601 Fishers Lane, Room 6B51 
Bethesda, MD, 20892-9827 
Tel:  240-627-3515 
Fax:  301-480-4258 
Email: jlaurienzo@niaid.nih.gov 
ITN Clinical Operations Manager 
Emerson Sosa 
Clinical Operations Manager 
Clinical Trials Group 
Immune Tolerance Network 
University of California, San Francisco 
Diabetes Center 
513 Parnassus Avenue, HSW 11, Box 0534 
San Francisco, CA 94143-0534Phone: 415-218-7981 
Email:  
esosa@immunetolerance.org 
This clinical study is supported and conducted by the Immune Tolerance Network, which is sponsored by 
the National Institute of Allergy and Infectious Diseases. 
This document is confidential. It is provided for review only to investigators, potential investigators, 
consultants, study staff, and applicable independent ethics committees or institutional review boards. It is 
understood that the contents of this document will not be disclosed to others without written authorization 
from ITN and NIAID unless it is necessary to obtain informed consent from potential study participants. 
 
 
 

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INVESTIGATOR SIGNATURE PAGE  
Trial ID:  ITN070AD 
Protocol Version: 7.0 
 
Date:  10 December 2021 
IND # Not applicable 
Protocol Chair:  Gideon Lack, MD 
Title:    Follow up of LEAP Participants and Their Families: LEAP Trio 
Return Signed Form to:  
The original signature page must be kept for your records. Return an electronic PDF copy of the 
signed signature page to the DAIT Regulatory Management Center via the applicable DAIT RMC 
email address for the protocol/network.  
DAITRegistration_SiteRegistration@ppdi.com 
I confirm that I have read the above protocol in the latest version. I understand it, and I will 
work according to the principles of good clinical practice (GCP) as described in the US Code of 
Federal Regulations (CFR) on the Protection of Human Subjects —45 CFR part 46 and 21 CFR parts 
50, 56, and in the International Conference on Harmonization (ICH) document entitled Integrated 
Addendum to ICH E6(R1): Guideline for Good Clinical Practice E6(R2). Further, I will conduct the 
study in keeping with local legal and regulatory requirements. 
 
As the principal investigator, I agree to carry out the study by the criteria written in the protocol 
and understand that no changes can be made to this protocol without written permission of the 
NIAID.  
 
*The site Principal Investigator should sign and date at the indicated location below. A written 
signature/date is acceptable (e.g., scanned and sent via email as a PDF version). An electronic 
signature is also acceptable (e.g., sent via email as a PDF version). 
 
 
__________________________________________                                        
Principal Investigator                        (Print)                     
 
 
__________________________________________         _________________ 
Principal Investigator                        (Sign)                      Date 
 
 
 
 
 

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TABLE OF CONTENTS 
INVESTIGATOR SIGNATURE PAGE ...................................................................................... 3 
TABLE OF CONTENTS ............................................................................................................. 4 
SYNOPSIS .................................................................................................................................. 9 
ABBREVIATIONS.................................................................................................................... 12 
1. 
BACKGROUND AND RATIONALE ................................................................................ 13 
1.1 
INITIAL STUDIES IN INFANTS AT HIGH RISK FOR ALLERGY ................................. 13 
1.2 
RATIONALE FOR LONG-TERM FOLLOW-UP OF LEAP PARTICIPANTS DURING AD 
LIBITUM PEANUT CONSUMPTION ............................................................................ 14 
1.3 
RATIONALE FOR THE STUDY OF LEAP SIBLINGS POPULATION: ASSESSING RISK FOR 
ALLERGIC SENSITIZATION ....................................................................................... 15 
1.3.1 
Anecdotal Observations from LEAP Families .......................................................................... 15 
1.3.2 
Questionnaires to LEAP Families ............................................................................................. 15 
1.3.3 
Plans for Current Study of Siblings ........................................................................................... 16 
1.4 
RATIONALE FOR GENETIC STUDIES IN LEAP PARTICIPANTS AND FAMILIES, 
INCLUDING PARENTS ................................................................................................ 16 
1.5 
SUMMARY OF KNOWN AND POTENTIAL RISKS AND BENEFITS FOR PARTICIPANTS
 ..................................................................................................................................... 17 
2. 
OBJECTIVES ................................................................................................................... 17 
2.1 
CO-PRIMARY OBJECTIVES ......................................................................................... 17 
2.2 
SECONDARY OBJECTIVES.......................................................................................... 17 
2.3 
EXPLORATORY OBJECTIVES ..................................................................................... 18 
3. 
STUDY DESIGN ............................................................................................................... 19 
3.1 
DESCRIPTION .............................................................................................................. 19 
3.2 
STUDY ENDPOINTS ..................................................................................................... 21 
3.2.1 
Co-Primary Endpoints ............................................................................................................... 21 
3.2.2 
Secondary Endpoints ................................................................................................................. 21 
3.2.3 
Exploratory Endpoints .............................................................................................................. 22 
3.3 
STUDY DEFINITIONS .................................................................................................. 22 
3.3.1 
Determination of Primary Peanut Allergy ................................................................................. 22 
3.3.2 
Determination of Peanut Sensitization ...................................................................................... 22 
3.3.3 
Definition of Rhinoconjunctivitis .............................................................................................. 23 
3.3.4 
Definition of Asthma ................................................................................................................. 23 
3.3.5 
Definition of Peanut Oral Allergy Syndrome ............................................................................ 23 
3.4 
DATA AND SAFETY MONITORING AND STOPPING RULES ..................................... 24 
3.4.1 
Ongoing Review........................................................................................................................ 24 
3.4.2 
Stopping Rules Guidance .......................................................................................................... 24 

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3.5 
STUDY DURATION ...................................................................................................... 24 
4. 
ELIGIBILITY ................................................................................................................... 25 
4.1 
LEAP PARTICIPANTS .................................................................................................. 25 
4.1.1 
Inclusion Criteria – LEAP Participants ..................................................................................... 25 
4.1.2 
Exclusion Criteria – LEAP Participants .................................................................................... 25 
4.2 
LEAP SIBLINGS ............................................................................................................ 25 
4.2.1 
Inclusion Criteria – LEAP Siblings ........................................................................................... 25 
4.2.2 
Exclusion Criteria – LEAP Siblings .......................................................................................... 25 
4.3 
LEAP PARENTS ............................................................................................................ 25 
4.3.1 
Inclusion Criteria – LEAP Parents ............................................................................................ 25 
4.3.2 
Exclusion Criteria – LEAP Parents ........................................................................................... 26 
4.4 
PREMATURE TERMINATION OF A PARTICIPANT FROM THE STUDY .................... 26 
5. 
STUDY THERAPIES AND MEDICATIONS .................................................................... 26 
6. 
STUDY ASSESSMENTS AND WINDOWS ...................................................................... 26 
6.1 
VISIT WINDOWS .......................................................................................................... 26 
6.1.1 
Scheduled Visits ........................................................................................................................ 26 
6.1.2 
Window to Complete Visit 144 ................................................................................................. 26 
6.1.3 
Definition of Enrollment and Study Participation ..................................................................... 26 
6.2 
VISIT TYPES ................................................................................................................. 27 
6.2.1 
Clinic ......................................................................................................................................... 27 
6.2.2 
Home ......................................................................................................................................... 27 
6.2.3 
Telephone .................................................................................................................................. 27 
6.3 
GENERAL ASSESSMENTS ........................................................................................... 28 
6.3.1 
Non-dietary Assessments .......................................................................................................... 28 
6.3.2 
Dietary Assessments ................................................................................................................. 29 
6.4 
CLINICAL LABORATORY ASSESSMENTS ................................................................. 29 
6.5 
ALLERGY ASSESSMENTS ........................................................................................... 29 
6.6 
SKIN-PRICK TESTING .................................................................................................. 29 
6.7 
PEANUT CHALLENGES ............................................................................................... 30 
6.7.1 
Conduct ..................................................................................................................................... 30 
6.7.2 
Outcome .................................................................................................................................... 34 
7. 
TOLERANCE ASSAYS .................................................................................................... 35 
7.1 
MECHANISTIC HYPOTHESES ..................................................................................... 35 
7.2 
PROPOSED MECHANISTIC ASSAYS ........................................................................... 36 
7.2.1 
Serum/Plasma Assays ............................................................................................................... 36 
7.2.2 
Cellular Assays.......................................................................................................................... 37 
7.2.3 
DNA Assays .............................................................................................................................. 38 

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7.2.4 
Gene Expression Profiling ........................................................................................................ 38 
7.3 
FUTURE AND UNPLANNED STUDIES ........................................................................ 38 
8. 
ADVERSE EVENTS ......................................................................................................... 39 
8.1 
OVERVIEW ................................................................................................................... 39 
8.2 
DEFINITIONS ............................................................................................................... 39 
8.2.1 
Adverse Event ........................................................................................................................... 39 
8.2.2 
Serious Adverse Event .............................................................................................................. 39 
8.3 
COLLECTING AND RECORDING ADVERSE EVENTS ................................................ 40 
8.3.1 
Collection Period ....................................................................................................................... 40 
8.3.2 
Collecting Adverse Events ........................................................................................................ 40 
8.3.3 
Recording Adverse Events ........................................................................................................ 40 
8.3.4 
Recording Serious Adverse Events ........................................................................................... 40 
8.4 
GRADING AND ATTRIBUTION OF ADVERSE EVENTS ............................................. 41 
8.4.1 
Grading Criteria ........................................................................................................................ 41 
8.4.2 
Attribution ................................................................................................................................. 41 
8.5 
REPORTING SERIOUS ADVERSE EVENTS ................................................................. 41 
8.5.1 
Reporting SAEs to the Sponsor ................................................................................................. 41 
8.5.2 
Reporting SAEs to the DSMB .................................................................................................. 42 
8.5.3 
Reporting SAEs and Unanticipated Problems to the Ethics Committee ................................... 42 
9. 
STATISTICAL CONSIDERATIONS AND ANALYTICAL PLAN ................................... 42 
9.1 
ANALYSIS SAMPLES ................................................................................................... 42 
9.2 
ANALYSIS OF ENDPOINTS ......................................................................................... 43 
9.2.1 
Primary Endpoint ...................................................................................................................... 43 
9.2.2 
Secondary and Exploratory Endpoints ...................................................................................... 44 
9.2.3 
Safety Analysis.......................................................................................................................... 46 
9.2.4 
Relevant Medical History.......................................................................................................... 46 
9.2.5 
Use of Medications ................................................................................................................... 46 
9.3 
SAMPLE SIZE ............................................................................................................... 46 
9.4 
MISSING DATA ............................................................................................................ 49 
9.5 
REPORTING DEVIATIONS FROM THE ORIGINAL STATISTICAL PLAN ................... 49 
10. ACCESS TO SOURCE DATA/DOCUMENTS .................................................................. 49 
11. QUALITY CONTROL AND QUALITY ASSURANCE .................................................... 50 
12. ETHICAL CONSIDERATIONS AND COMPLIANCE WITH GOOD CLINICAL PRACTICE
 ..................................................................................................................................... 50 
12.1 
STATEMENT OF COMPLIANCE .................................................................................. 50 
12.2 
INFORMED CONSENT ................................................................................................. 51 
12.3 
PRIVACY AND CONFIDENTIALITY ............................................................................ 51 

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13. PUBLICATION POLICY ................................................................................................. 51 
14. REFERENCES ................................................................................................................. 52 
APPENDIX 1. SCHEDULE OF EVENTS ................................................................................. 53 
APPENDIX 2. PREVIOUSLY USED ALGORITHM FOR DETERMINING PEANUT ALLERGY 
STATUS ....................................................................................................................... 55 
 

 
 
 
Protocol ITN070AD 
v 5.0 
5 November 2020 
LEAP Trio, IRAS ID 195451 
Immune  
Tolerance 
Network 
 
 
LIST OF FIGURES 
Figure 1. LEAP Screening, Participant and Sibling Cohorts ................................................................... 19 
Figure 2. Dates and ages in months during participation in phases of LEAP and related studies ........... 21 
Figure 3. Power estimates for allergy rates ............................................................................................. 47 
Figure 4. Power estimates for sensitization rates ..................................................................................... 49 
Figure 5. Prior Algorithm for Determination of peanut allergy in individuals who do not undergo an oral 
food challenge………………………………………………………………………………….58 
 
 
 
LIST OF TABLES 
Table 1. Criteria for determining the outcome of food challenge ............................................................ 35 
Table 2. Attribution of Adverse Events ................................................................................................... 41 
 
LIST OF APPENDICES 
APPENDIX 1. Schedule of Events .......................................................................................................... 53 
APPENDIX 2. Previously Used Algorithm For Determining Peanut Allergy Status ............................. 55 
 
 
 
 

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SYNOPSIS 
Title 
Follow up of LEAP Participants and Their Families: LEAP Trio  
Sponsor 
 
Legal 
Representative in 
UK 
The National Institute of Allergy & Infectious Diseases, US 
 
 
Clinical Technology Centre (International) Limited, Cambridge, 
United Kingdom 
Conducted by 
Immune Tolerance Network 
Protocol Chair  
Gideon Lack, MD 
Study Design 
This is a long-term assessment of the LEAP randomized controlled 
study of early peanut consumption. LEAP participants are followed 
during an extended period of ad-libitum peanut consumption and then 
assessed for peanut allergy and other allergic outcomes at 
approximately age 12. In addition, siblings and parents will be 
assigned to the intervention or control group based on the prior 
randomization of their LEAP participant sibling or child, respectively. 
Co-Primary 
Objectives 
1. To assess whether early consumption of peanuts by high-risk 
infants results in a decreased risk of peanut allergy in children 
approximately 12 years of age. 
2. To determine the prevalence of sensitization in younger siblings 
of LEAP participants who resided in the home at the time of the 
LEAP intervention, comparing younger siblings of LEAP 
participants who consumed peanut to younger siblings of LEAP 
participants who avoided peanut. 
Co-Primary 
Endpoint 
LEAP Participants 
The primary endpoint is the rate of peanut allergy in LEAP 
participants at 144 months of age. The strategy for determination of 
peanut allergy is outlined in section 3.3. 
LEAP Siblings 
The primary endpoint is the rate of peanut sensitization in younger 
siblings who resided in the home of the LEAP participant on or before 
LEAP Visit 60, as assessed in the current LEAP Trio study. The 
strategy for determination of peanut sensitization is outlined in section 
3.3.2. 

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Secondary 
Endpoints 
LEAP Participants, Siblings, and Parents: Clinical 
1. Amount of peanut consumption as measured by peanut 
consumption questionnaires.  
2. Skin-prick test wheal sizes to peanut, other select foods, and 
aeroallergens.  
3. Specific-IgE measurements to peanut, other select foods, and 
aeroallergens. 
4. Eczema severity by clinical assessment of SCORAD in LEAP 
participants and siblings. 
5. Prevalence of asthma, rhinitis and eczema evaluated using a 
combination of questionnaires, examinations, and lung function 
testing in LEAP participants and siblings.  
6. Prevalence of asthma, rhinitis and eczema evaluated using 
questionnaires in parents. 
7. Prevalence of peanut allergy in LEAP siblings and parents. 
8. Peanut-related adverse events in LEAP participants and siblings.  
Exploratory 
Endpoints 
LEAP Participants, Siblings, and Parents 
1. Prevalence of allergies (other than peanut). 
2. Dietary assessment questionnaires in LEAP participants. 
3. Transepidermal Water Loss (TEWL) in LEAP participants and 
siblings. 
4. Household peanut consumption questionnaires and concentration 
of environmental peanut protein in dust collected from the 
participants’ homes. 
 
LEAP Participants, Siblings, and Parents: Mechanistic 
Endpoints for mechanistic studies are described in section 7. 
Inclusion 
Criteria –  
LEAP 
Participants 
1. Participation in LEAP. 
2. Age at least 114 months (9.5 years). 
3. Willingness to participate in at least one study data collection (i.e. 
questionnaire, skin prick testing or blood draw) procedure. 
4. Assent by child and informed consent by parent or legal guardian.  
For participants only returning a Questionnaire in any format, 
assent/informed consent will be implied. For participants only 
completing a telephone visit, assent/informed consent will be given 
verbally. 
Exclusion 
Criteria  –  
LEAP 
Participants 
There are no exclusion criteria for LEAP participants. 
Inclusion 
Criteria –  
LEAP Siblings 
1. Sibling of LEAP participant. 
2. Willingness to participate in at least one study data collection (i.e. 
questionnaire, skin prick testing or blood draw). 

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3. Assent by child and informed consent by parent or legal guardian 
if child is younger than 16; siblings aged 16 and over will provide 
their own consent. For participants only returning a Questionnaire 
in any format, assent/informed consent will be implied). For 
participants only completing a telephone visit, assent/informed 
consent will be given verbally. 
Exclusion 
Criteria –   
LEAP Siblings 
There are no exclusion criteria for LEAP siblings. 
Inclusion 
Criteria –  
LEAP Parents 
1. Biological parent of LEAP participant.  
2. Willingness to participate in at least one study data collection (i.e. 
questionnaire, skin pricking testing or blood draw). 
3. Informed consent. For participants only returning a Questionnaire 
in any format, informed consent will be implied. For participants 
only completing a telephone visit, informed consent will be given 
verbally. 
Exclusion 
Criteria –  
LEAP Parents 
There are no exclusion criteria for LEAP parents. 
 
 
 

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ABBREVIATIONS 
AE 
adverse event 
AD 
atopic dermatitis 
AUC 
area under the curve 
CBC 
complete blood count 
CFR 
Code of Federal Regulations 
CRD 
component resolved diagnosis 
CRF 
case report form 
CRO 
contract research organization 
DAIT 
Division of Allergy, Immunology, & Transplantation 
DSMB 
Data and Safety Monitoring Board 
FA 
food allergy 
FDA 
US Food and Drug Administration  
FFQ 
Food Frequency Questionnaire 
FLG 
filaggrin 
GCP 
good clinical practice 
ICH 
International Conference on Harmonisation 
ITN 
Immune Tolerance Network 
MedDRA 
Medical Dictionary for Regulatory Activities 
NCI-CTCAE 
National Cancer Institute Common Terminology Criteria 
for Adverse Events 4.03 (published June 10, 2010) 
NIAID 
National Institute of Allergy and Infectious Diseases 
NIH 
National Institutes of Health 
OAS 
Oral Allergy Syndrome 
PA 
peanut allergy 
PD 
pharmacodynamics 
PK 
pharmacokinetics 
SACCC 
Statistical and Clinical Coordinating Center 
SAE 
serious adverse event 
SAP 
statistical analysis plan 
SCORAD 
SCORing Atopic Dermatitis 
TEWL 
transepidermal water loss 
WAO 
World Allergy Organization 

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WHO 
World Health Organization 
 
1. 
BACKGROUND AND RATIONALE 
1.1 
INITIAL STUDIES IN INFANTS AT HIGH RISK FOR ALLERGY 
Food allergies and peanut allergy (PA) are becoming increasingly common conditions 
and are now an important public health concern.1 Dietary avoidance of peanut in early 
life has been recommended in many countries.2 However, there is evidence that the 
prevalence of PA is lower in countries such as Israel where children are fed peanut from 
an early age. The NIAID-funded LEAP (Learning Early About Peanut allergy) Study 
(Protocol ITN032AD, NCT00329784) recently demonstrated that the early consumption 
of peanut in high-risk infants successfully reduced the prevalence of peanut allergy at 
five years of age when compared to peanut avoidance (81% relative reduction, intention 
to treat analysis).3 The EAT (Enquiring About Tolerance) Study demonstrated that peanut 
and egg consumption during infancy reduced the rates of allergy to these allergens at 3 
years of age, in a general population of infants, within the per-protocol population.4 
The LEAP-On Study (Protocol ITN049AD, NCT01366846) was a follow-on study to 
LEAP and investigated whether children who consumed peanut remained protected 
against developing peanut allergy even after cessation of peanut consumption for a period 
of 12 months. A total of 556 participants (88.5% (556/628)) from the LEAP trial were 
enrolled in the follow-on study and the rate of adherence to avoidance was high (90.4% 
in the peanut-avoidance group, 69.3% in the peanut-consumption group). At 72 months, 
peanut allergy remained significantly higher in the LEAP peanut-avoidance group 
compared to the LEAP peanut-consumption group, 18.6% vs 4.8% (p<0.001) 
respectively.5 Three new cases of allergy developed in each group, but after 12 months of 
avoidance there was no significant increase in the prevalence of allergy among 
participants in the consumption group (3.6% [10 of 274 participants] at 60 months and 
4.8% [13 of 270] at 72 months, P = 0.25). These clinical findings were associated with 
immunological changes (levels of Ara h2-specific peanut IgE, peanut specific IgE and 
IgG4 levels) suggesting immune tolerance. The key finding of the LEAP studies is that 
early introduction and consumption of peanut until 60 months of age causes a reduction 
in peanut allergy that persists at 72 months of age, even with a 12-month period of 
avoidance. 
A follow up study of LEAP participants, and studies of their siblings and parents, are 
now warranted to determine if the effects of early tolerance persist to 12 years of age, to 
assess the impact of household exposure to peanut among siblings of LEAP participants, 
and to examine parental characteristics that may impact the development of food allergy. 
Children who participated in LEAP (and LEAP-On), any siblings, and both parents will 
be asked to participate in one of three research studies which together will be referred to 
as the “LEAP Trio” (Protocol ITN070AD): 
• 
The LEAP Ad Lib Study will enroll original LEAP participants. 
• 
The LEAP Siblings Study will enroll siblings of LEAP participants. 
• 
The LEAP Parents Study will enroll the parents of LEAP participants. 
 

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Rationale for the study of these three separate groups is provided in following sections. 
1.2 
RATIONALE FOR LONG-TERM FOLLOW-UP OF LEAP 
PARTICIPANTS DURING AD LIBITUM PEANUT CONSUMPTION 
The LEAP study findings have been adopted by international societies, including 
Allergy, Pediatric and Dermatology Societies. In January, 2017 NIAID published 
Addendum Guidelines for the Prevention of Peanut Allergy in the United States.6 It is 
therefore becoming increasingly important to fully understand the long term clinical and 
nutritional outcomes of the LEAP Study beyond the 12 months study period of the 
LEAP-On Study and with participants having eaten peanut ad-libitum. The LEAP 
Participant cohort will be rigorously assessed for peanut allergy at 12 years of age to 
determine if the LEAP Study intervention achieved tolerance that is independent of 
ongoing peanut consumption at 12 years of age as opposed to transient desensitization 
that is dependent on ongoing peanut ingestion. 
The LEAP-On Study demonstrated that for the peanut consumption group, their non-
allergic status remained stable over 12 months of peanut avoidance.5 If the early-life 
dietary intervention in LEAP (i.e. peanut consumption) does induce a long-term effect on 
the natural history of peanut allergy, the most likely mechanism is one involving a long-
term alteration in the morphology and function of the developing immune system. This 
will be further investigated in this study.  
In LEAP-On, participants with peanut allergy at 72 months had higher levels of Ara h2-
specific IgE compared to those who did not have peanut allergy.5 The mean level of Ara-
h2-specific IgE, which had declined significantly in the peanut consumption group in the 
LEAP trial (p<0.001) remained low at 72 months, after 12 months of peanut avoidance. 
By contrast, the mean levels of Ara-h2-specific IgE in the peanut avoidance group were 
significantly higher at 60 months and 72 months compared to the peanut consumption 
group (p<0.001).  
When considering IgG4 and the ratio of peanut specific IgG4:IgE, they were both 
significantly higher in the peanut consumption group compared to the peanut avoidance 
group (p<0.001) at 60 months but also 72 months (after a yearlong period of avoidance 
(p<0.001)). These findings therefore favour those parameters traditionally associated with 
tolerance. However, in the consumption group, values of peanut IgG4 had started to 
decrease, even at 30 months of age (when peanut consumption was ongoing). The longer-
term trajectory of IgG4, and clinical relevance thereof, will become apparent after the 
determination of peanut allergy at 12 years of age in the LEAP Participant cohort. 
Many children with peanut allergy develop concomitant sensitization and allergy to other 
peanut-related foods such as tree nuts and sesame. This adds an additional burden both in 
terms of quality of life and morbidity associated with living with allergic disease. 
Similarly, early life eczema and egg allergy have been shown to be a risk for asthma, and 
possibly allergic rhino-conjunctivitis. We will assess the natural history of these 
conditions in an atopic population and will assess whether the benefits of the LEAP 
intervention extend to other atopic disease and other food allergens. 

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The nutritional consequences of consuming peanut in the LEAP Study has been 
published.7 The current study will help determine if these benefits are sustained beyond 
the first six years of life and after participants have been eating peanut ad-libitum. 
We therefore aim to further investigate the clinical, immunological and nutritional safety 
findings of the LEAP and LEAP-On Studies. The findings will provide an opportunity to 
improve our understanding of how early-life introduction of peanuts may promote the 
development of tolerance at 12 years of age and will serve as an additional safety 
evaluation of this nutritional intervention. 
1.3 
RATIONALE FOR THE STUDY OF LEAP SIBLINGS POPULATION: 
ASSESSING RISK FOR ALLERGIC SENSITIZATION  
1.3.1 Anecdotal Observations from LEAP Families 
During the LEAP Study, from discussions with families at the time of scheduled visits 
and from observations made during study-related phone calls, it became apparent that 
that siblings, especially younger ones, of LEAP participants born during LEAP may have 
been disproportionately affected with suspected peanut allergy. The initial analysis 
suggested that 13 siblings of 319 LEAP consumers may have developed peanut allergy as 
compared to none of the siblings born to 320 LEAP avoiders. This analysis was based on 
allergic symptoms after ingestion volunteered by parents and not by systematic 
questioning or formal allergy testing. 
This finding could have reflected either a real difference in the prevalence of peanut 
allergy in the two groups or reporting bias on the part of parents in households where 
peanut consumption - and opportunity for exposure - was high. However, it raised the 
concern that intentional peanut consumption as specified for consuming children in the 
LEAP study could increase the risk of peanut allergy among siblings in the same 
household. This concern was discussed by the LEAP Study Management Team and 
independent ethical advice was sought. 
1.3.2 Questionnaires to LEAP Families 
To further investigate the anecdotal observations, local Ethics Committee approval for 
issuing a questionnaire (the ‘LEAP and LEAP-On Sibling Peanut Allergy Questionnaire’) 
was obtained in October 2013 through amendments to the LEAP Study (04/Q0403/13) 
and the LEAP-On Study (10/H0711/77). All LEAP participant families were then 
informed of the background to and need for completion of the LEAP and LEAP-On 
Sibling Peanut Allergy Questionnaire. Out of 640 families, 619 (97%) were contacted 
and of these, 600 (94%) provided informed consent and completed the questionnaire. 
Nineteen declined informed consent. 523 families had at least one more child in addition 
to the LEAP participant while 77 families did not have a LEAP sibling. In total, these 
consented families provided information for 746 siblings. 
The LEAP and LEAP-On Sibling Peanut Allergy Questionnaire revealed a questionnaire-
based diagnosis of peanut allergy of 4.2% in younger siblings of the LEAP avoidance 
group participants and 9.2% in younger siblings of the LEAP consumption group 
participants (p-value of difference in proportions is 0.057 using a one-tailed Wald 
analysis). These findings therefore supported the original observation that early peanut 
introduction in high-risk children may have led to an increase in the rates of peanut 

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allergy in the younger siblings of children who consumed peanut on the LEAP Study as 
compared to the younger siblings of LEAP participants who avoided peanut. However, 
similar to the anecdotal observations from LEAP families, these questionnaire-based 
findings may also have arisen due to ascertainment bias because the siblings of LEAP 
peanut avoiders are less likely to have been exposed to peanut and therefore have had less 
opportunity to manifest symptoms of peanut allergy. An additional source of uncertainty 
arises from the 78 younger siblings (56 in the avoidance group and 22 in the consumption 
group) for whom the questionnaire result was a ‘don’t know’ outcome due to the fact that 
the sibling may never have eaten peanut or ever undergone peanut allergy testing. 
1.3.3 Plans for Current Study of Siblings 
The LEAP Siblings cohort in the current study therefore aims to further investigate the 
findings of the LEAP and LEAP-On Sibling Peanut Allergy Questionnaire using a more 
rigorous scientific methodology to exclude the possibility of ascertainment bias. The 
findings will serve as an important safety evaluation. Study of this cohort will also 
provide an opportunity to improve our understanding of how early introduction of 
peanuts promotes the development of tolerance, and will estimate the prevalence of 
peanut allergy in siblings of LEAP participants. 
1.4 
RATIONALE FOR GENETIC STUDIES IN LEAP PARTICIPANTS AND 
FAMILIES, INCLUDING PARENTS 
There are many risk factors associated with the development of food allergy (FA), 
including atopic family history, male sex (at least in childhood), ethnicity, atopic 
dermatitis (AD), and related genetic polymorphisms. Although genetic factors are clearly 
important in the development of food allergy, the increase in prevalence of food allergy 
has occurred over a short period of human evolution, implying that food allergy does not 
arise as a result of germline genetic changes alone. It therefore seems plausible that one 
or more environmental exposures can induce epigenetic changes that interrupt the default 
immunologic state of tolerance to foods. 
There are however many studies to suggest that genetic factors play a role in the 
development of food allergy. For example, a family history of FA is itself a risk factor for 
FA. A child has a 7-fold increase in the risk of peanut allergy if there is a parent or 
sibling with PA.8  
The complex interplay between genetic and environmental factors giving rise to FA is 
perhaps best demonstrated by comparing concordance rates for allergy between 
genetically identical (monozygotic) and nonidentical (dizygotic) subjects.  
Although previous twin studies have estimated a high degree of heritability for atopic 
diseases, such as asthma (87%) and AD (74%), a study by Sicherer et al.9 of 58 twin pairs 
estimated the heritability for PA to be as high as 82% to 87%. In a recent review, Hong et 
al.10 highlighted more than 10 genes (several involved in allergen presentation, a Th2-
skewed immune system, or both) that have been associated with FA or food sensitization. 
However, genetically determined skin barrier dysfunction—associated with mutations in 
the gene encoding filaggrin (FLG)—has attracted the most interest, as this is known to 
predispose to multiple systemic atopic diseases. 

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The LEAP Study families represent an ideal opportunity to assess for genetic influences 
on peanut allergy in an atopic, well-phenotyped population. The mechanistic relevance of 
those assessments of various LEAP populations is that we would expect all study 
participants who develop peanut allergy to have filaggrin null mutations, or possibly 
mutations in other genes that encode for proteins that are important in skin barrier 
function. A variety of analysis approaches may be taken. Some of these are illustrated in 
a recent review of the influence of genetics on allergic outcomes.11 
1.5 
SUMMARY OF KNOWN AND POTENTIAL RISKS AND BENEFITS FOR 
PARTICIPANTS  
The main risks of study participation relate to study assessments. These include 
venipuncture, skin prick testing and oral food challenge. Venipuncture is associated with 
a moderate risk of discomfort and a small risk of hematoma. Skin prick testing may be 
associated with localized swelling and redness. 
Oral peanut food challenge in non-allergic subjects does not carry risk. In the subset of 
allergic individuals, risks associated with peanut challenges include nausea, vomiting, 
itching, urticaria, angioedema, asthma, other respiratory symptoms, and anaphylaxis, 
which can be life threatening. 
In the study, DNA will be collected for genetic analyses. This is associated with a risk of 
loss of privacy. 
2. 
OBJECTIVES 
2.1 
CO-PRIMARY OBJECTIVES 
1. To assess whether early consumption of peanuts by high-risk infants results in a 
decreased risk of peanut allergy in children approximately 12 years of age. 
2. To determine the prevalence of sensitization in younger siblings of LEAP 
participants who resided in the home at the time of the LEAP intervention, 
comparing younger siblings of LEAP participants who consumed peanut to 
younger siblings of LEAP participants who avoided peanut. 
2.2 
SECONDARY OBJECTIVES 
 
LEAP Participants 
1. 
To assess the effect of peanut consumption or avoidance in an at-risk study 
population (former LEAP participants) on additional allergic disease outcomes such 
as:  a) allergic sensitization to select ingested allergens, b) allergic sensitization to 
select aeroallergens, c) reported type 1 immediate-onset food allergy, d) seasonal 
and perennial rhino-conjunctivitis, e) oral allergy syndrome, and f) asthma. 
2. 
To assess the long-term safety of the LEAP intervention as determined by the 
frequency and severity of peanut-related adverse events as well as by nutritional 
evaluations. 
3. 
To assess eczema severity by clinical assessment of SCORAD. 
4. 
To define the long-term immunological impact of the intervention using cellular 
and humoral assessments of immune response related to the development of allergy 
or tolerance to specific allergens.  

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 LEAP Siblings  
1. 
To assess the impact of environmental peanut exposure, derived from LEAP Food 
Frequency Questionnaires (FFQs) and peanut levels in bed dust at the LEAP 60 
month visit, on prevalence of peanut sensitization and allergy in siblings of LEAP 
participants.  
2. 
To control for allergen-specific effect of environmental exposure by assessing the 
prevalence of sensitization and reported food allergy to common food allergens 
including hen's egg white, cow's milk, sesame seed, and the five tree nuts assessed 
in the  LEAP Study (Brazil nut, hazel nut, cashew, walnut, almond) in siblings of 
LEAP participants.  
3. 
To assess the impact of peanut consumption in siblings during the first and second 
years of life on the development of peanut sensitization and allergy in these 
siblings of LEAP participants. 
4. 
To characterize the allergy status in all siblings of LEAP participants. 
5. 
To assess eczema severity by clinical assessment of SCORAD in siblings of LEAP 
participants. 
6. 
To assess seasonal and perennial rhinoconjunctivitis, asthma, and eczema in the 
siblings of LEAP participants. 
7. 
To assess safety as determined by the frequency and severity of peanut-related 
adverse events in the siblings of LEAP participants. 
LEAP Parents  
1. 
To characterize the allergy status in all parents of LEAP participants.  
2. 
To characterize environmental peanut exposure derived from peanut consumption 
questionnaires in all parents of LEAP participants. 
3. 
To assess the prevalence of sensitization to common food and aeroallergens in all 
parents of LEAP participants. 
4. 
To assess seasonal and perennial rhinoconjunctivitis, asthma, and eczema in all 
parents of LEAP participants. 
2.3 
EXPLORATORY OBJECTIVES 
1. 
To assess skin barrier function by clinical assessment of Trans Epidermal Water 
Loss (TEWL) measurement in LEAP participants and siblings of LEAP 
participants. 
2. 
To assess mechanistic correlates of allergy and tolerance, as described in section 7.  
3. 
To collect genetic material for possible future research. 
4. 
To characterize household peanut consumption and concentration of environmental 
peanut protein in dust. 
5. 
To assess the impact of peanut consumption on dietary assessments derived from 3-
day food diaries in LEAP participants. 
6. 
To summarize allergies to foods other than peanut. 

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3. 
STUDY DESIGN 
3.1 
DESCRIPTION 
This is a long-term assessment of the LEAP randomized controlled study of early peanut 
consumption. LEAP participants are followed during an extended period of ad-libitum 
peanut consumption and then assessed for peanut allergy and other allergic outcomes at 
approximately age 12. In addition siblings and parents will be assigned to the intervention 
or control group based on the prior randomization of their LEAP participant sibling or 
child, respectively. 
The flow of LEAP participants from screening through LEAP and LEAP-On is shown in 
Figure 1. The figure indicates how many individuals are likely to be available from 
various cohorts for enrollment into the study. In addition, siblings and parents of LEAP 
participants will be followed for allergic and genetic parameters. 
899 screened for 
LEAP
640
LEAP Trial cohort
2829 children’s families
made contact with 
LEAP study team
2006-2009
1930 not screened
1701 declined
198 ineligible for screening
31 screening cancelled
76
Group IV
SPT>4mm
65 excluded
26 cohort closed
13 withdrew consent
26 other
272+47 = 319 
Assigned to 
Consumption
270+51 = 321
Assigned to 
Avoidance
118
Group I
Eczema not severe
834
LEAP 
Screening Cohort
Enrolled in 
LEAP?
No
No
Yes
314 Avoiders were 
included in LEAP ITT 
analysis and allowed 
to enroll in LEAP-On
7 had missing allergy 
outcomes
314 Consumers were 
included in LEAP ITT 
analysis and allowed 
to enroll in LEAP-On
5 had missing 
allergy outcomes
316 Avoiders 
potentially eligible for 
LEAP Trio study
5 families declined to 
be contacted
314 Consumers 
potentially eligible for 
LEAP Trio Study
5 families declined to 
be contacted
282 Avoiders 
enrolled in 
LEAP-On
274 Consumers 
enrolled in 
LEAP-On
556
LEAP-On Cohort
Potential LEAP Trio Cohorts
630 LEAP Participants
 945 LEAP Parents
746 LEAP Siblings
TOTAL 2321
222 younger sibs
153 older sibs in same 
household
474 parents
207 younger sibs
164 older sibs in same 
household
471 parents
Avoid or 
Consume
Avoid
Ad Lib
 
Figure 1. LEAP Screening, Participant, Sibling, and Parent Cohorts 
The number of siblings in the Ad Lib section are based on findings in a survey regarding siblings 
of LEAP participants. These numbers represent an estimate of the total participants expected to 
enroll. 

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Participants in the current study derive from participants in previous LEAP trials and 
their families. The LEAP Screening Cohort has been described.12 The LEAP study 
enrolled 640 participants,3 of whom 556 were also studied in the LEAP-On follow up 
study.5 All participants who enrolled in LEAP, including those who were not part of the 
intent-to-treat (ITT) population, are eligible to enroll in this study. Parents and siblings of 
LEAP participants who meet other eligibility criteria described in sections 4.2 and 4.3 are 
eligible to enroll. 
We reviewed the ages and dates of enrollment of participants in the LEAP trial. The 
times of enrollment and participation are shown in Figure 2. We assume the trial opens 
for enrollment in March 1, 2018. The last day for enrollment is August 31, 2022. The 
study was extended to accommodate for slowed enrollment due to the COVID-19 
pandemic. As a result of the study extension, participants are predicted to be older than 
originally planned. This difference is not likely to significantly alter the study outcomes, 
as allergic status is unlikely to change during this time interval. These parameters allow 
us to describe expected ages at enrollment of LEAP participants in the current study.  
Participant A, and similar participants who were relatively old at enrollment and enrolled 
relatively early in LEAP, could be as old as [10.9 months + (August 31, 2022 – February 
2, 2007)] = 332 + 5689 days or 16.5 years. 
For participant E, and similar participants who were relatively young at enrollment and 
enrolled relatively late in LEAP, the minimum age for eligibility applies (section 4.1.1). 
At the time of trial opening they would be as young as [4.5 months + (March 1, 2018 – 
March 20, 2009)] = 137 + 2844 days or 9.3 years. The minimum age of 9.5 years for 
eligibility means such participants could therefore be eligible for enrollment 0.2 years 
after March 1, 2018, or in May 2018 or after. 

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Figure 2. Dates and ages in months during participation in phases of LEAP and related studies 
The times for participation of LEAP participants in distinct phases of the LEAP studies are shown.  
 
3.2 
STUDY ENDPOINTS 
3.2.1 Co-Primary Endpoints 
LEAP Participants 
The primary endpoint is the rate of peanut allergy in LEAP participants at 144 months of 
age. The strategy for determination of peanut allergy is outlined in section 3.3. 
LEAP Siblings 
The primary endpoint is the rate of peanut sensitization in younger siblings who resided 
in the home of the LEAP participant on or before LEAP Visit 60, as assessed in the 
current LEAP Trio study. The strategy for determination of peanut sensitization is 
outlined in section 3.3.2. 
3.2.2 Secondary Endpoints 
LEAP Participants, Siblings, and Parents: Clinical 
1. Amount of peanut consumption as measured by peanut consumption questionnaires.  
2. Skin-prick test wheal sizes to peanut, other select foods, and aeroallergens.  
3. Specific-IgE measurements to peanut, other select foods, and aeroallergens. 
4. Eczema severity by clinical assessment of SCORAD in LEAP participants and 
siblings. 

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5. Prevalence of asthma, rhinitis and eczema evaluated using a combination of 
questionnaires, examinations, and lung function testing in LEAP participants and 
siblings.  
6. Prevalence of asthma, rhinitis and eczema evaluated using questionnaires in parents. 
7. Prevalence of peanut allergy in LEAP siblings and parents. 
8. Peanut-related adverse events in LEAP participants and siblings.  
3.2.3 Exploratory Endpoints 
LEAP Participants, Siblings, and Parents 
1. Prevalence of allergies (other than peanut). 
2. Dietary assessment questionnaires in LEAP participants. 
3. Transepidermal Water Loss (TEWL) in LEAP participants and siblings. 
4. Household peanut consumption questionnaires and concentration of environmental 
peanut protein in dust collected from the participants’ homes. 
LEAP Participants, Siblings, and Parents: Mechanistic 
Endpoints for mechanistic studies are described in section 7. 
3.3 
STUDY DEFINITIONS 
3.3.1 Determination of Primary Peanut Allergy 
Peanut allergy status will be determined by peanut challenge where possible. Peanut 
challenges are described in section 6.7. Participants meeting criteria for peanut oral 
allergy syndrome in the absence of primary peanut allergy as described in section 3.3.5 
will not be considered peanut allergic. 
Individuals consuming at least 2g of peanut protein without reaction on at least one 
occasion in the last year will be considered peanut tolerant.  
If oral challenge is not possible and participants do not meet the above criteria for 
tolerance, peanut allergy status will be determined by an internally and externally 
validated novel peanut allergy prediction model, the manuscript for which is currently in 
preparation: Developing a Prediction Model for Determination of Peanut Allergy Status 
in The LEAP Trio Study; Sever ML, Calatroni A, Du Toit G, et al. The model was 
developed using data from the LEAP participants, and incorporates peanut wheal size 
(mm), peanut specific IgE (kU/L), IgG4:IgE Ratio and Ara h2 (kU/L). The model 
performs with high level of accuracy, and continues to perform reasonably well in the 
context of missing blood or skin prick data. This model replaces the previously specified 
algorithm for determination of peanut allergy status. The details of the prior algorithm are 
included in the Appendix 2 for historical reference.  
3.3.2 Determination of Peanut Sensitization 
Peanut sensitization is defined as:  

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a) Sp-IgE ≥ 0.35kU/L or 
b) Sp-IgE to individual peanut component Ara h2 ≥ 0.1kU/L or  
c) Skin prick test to peanut ≥ 3mm. 
3.3.3 Definition of Rhinoconjunctivitis 
Perennial rhinoconjunctivitis. Sensitization to a perennial allergen and clinical history 
of rhinoconjunctivitis symptoms experienced when exposed to the relevant allergen. 
Seasonal rhinoconjunctivitis. Sensitization to a seasonal allergen and clinical history of 
rhinoconjunctivitis symptoms experienced during the relevant season. 
3.3.4 Definition of Asthma 
Asthma will be diagnosed if one of the following three definitions is met: 
• 
A history of cough, wheeze, or shortness of breath that meets all three of the 
following criteria: (1) was responsive to therapy with bronchodilators on two or more 
occasions in the previous 24 months, and (2) required one visit to a physician in the 
previous 24 months, and (3) occurred during the night, during early morning, or upon 
exercising in the intervals between exacerbations at any time in the previous 12 
months, or 
• 
Participant report of an asthma diagnosis on the Medical History CRF with current 
use of asthma medications (SABA or controller) on the Concomitant Medications 
CRF, or 
• 
Participant report of transient wheeze on exposure to a suspected allergen with 
confirmatory allergy tests. 
3.3.5 Definition of Peanut Oral Allergy Syndrome 
Peanut Oral Allergy Syndrome (OAS) will be diagnosed using the following criteria:  
An indeterminate or positive Visit 144 OFC with symptoms limited to typical 
OAS symptoms will be independently reviewed by allergists not participating in 
the OFC. A diagnosis of OAS without primary peanut allergy will made based on 
the following criteria: 
• 
OAS symptoms after eating an additional non-peanut food typically 
associated with OAS  
• 
Birch sensitisation on SPT (≥3 mm) or SpIgE (≥0.35 kU/L) 
• 
Ara h components consistent with diagnosis of OAS in the absence of 
primary peanut allergy as per the definition in section 3.3.1 
• 
No prior history of primary peanut allergy 
A negative Visit 144 OFC with symptoms limited to typical OAS symptoms will be 
considered OAS if the participant additionally meets the following criteria: 
• 
OAS symptoms after eating an additional non-peanut food typically 
associated with OAS  
• 
Birch sensitisation on SPT (≥3 mm) or SpIgE (≥0.35 kU/L) 

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• 
Ara h components consistent with diagnosis of OAS in the absence of 
primary peanut allergy as per the definition in section 3.3.1 
A participant that does not undergo an OFC, but is deemed not allergic as per the 
definition in section 3.3.1 will be considered to have OAS if the following criteria are 
met: 
• 
OAS symptoms after eating peanut and an additional non-peanut food 
typically associated with OAS  
• 
Birch sensitisation on SPT (≥3 mm) or SpIgE (≥0.35 kU/L) 
• 
Ara h components consistent with diagnosis of OAS in the absence of 
primary peanut allergy as per the definition in section 3.3.1 
 
3.4 
DATA AND SAFETY MONITORING AND STOPPING RULES 
3.4.1 Ongoing Review 
The protocol chair, the DAIT/NIAID medical monitor, the ITN clinical trial physician, 
and the NIAID Allergy and Asthma Data and Safety Monitoring Board (DSMB) will 
review safety data on an ongoing basis and at least annually. The DSMB may stop 
enrollment or participation in the trial at any time if it concludes that there are significant 
safety concerns.  
3.4.2 Stopping Rules Guidance 
3.4.2.1 Study-related Adverse Events  
Enrollment in the trial will be stopped pending review (1) if any death occurs or (2) if 
two participants are admitted to an intensive care unit for an adverse event related to 
study participation. 
Oral food challenges will be stopped pending review if two oral food challenges result in 
anaphylaxis associated with hypotension or hypoxia as defined in section 8.2.2 and which 
does not respond to two epinephrine injections. 
3.5 
STUDY DURATION 
This study will commence in 2018 with data collection continuing until  August 31, 2022. 
This will allow for the enrolment of former LEAP participants aged 12 years.  
The LEAP siblings and parents cohorts will be enrolled concurrently with the LEAP 
participant cohort.  
Study duration will be 234 weeks: 
• 
Enrollment phase will be 234 weeks (4 years, 6 months).  
• 
Study participation phase will be 1 visit.  

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4. 
ELIGIBILITY 
4.1 
LEAP PARTICIPANTS 
4.1.1 Inclusion Criteria – LEAP Participants 
Individuals must meet all of the following criteria to be eligible for this cohort in the 
study: 
1. Participation in LEAP. 
2. Age at least 114 months (9.5 years). 
3. Willingness to participate in at least one study data collection (i.e. questionnaire, skin 
pricking testing, or blood draw) procedure. 
4. Assent by child and informed consent by parent or legal guardian. For participants 
only returning a Questionnaire in any format, assent/informed consent will be 
implied. For participants only completing a telephone visit, assent/informed consent 
will be given verbally. 
4.1.2 Exclusion Criteria – LEAP Participants 
There are no exclusion criteria for LEAP Participants. 
4.2 
LEAP SIBLINGS 
4.2.1 Inclusion Criteria – LEAP Siblings 
Individuals must meet all of the following criteria to be eligible for this cohort in the 
study: 
1. Sibling of LEAP participant. 
2. Willingness to participate in at least one study data collection (i.e. questionnaire, skin 
pricking testing, or blood draw). 
3. Assent by child and informed consent by parent or legal guardian if child is younger 
than 16; siblings aged 16 and over will provide their own consent. For participants 
only returning a Questionnaire in any format, assent/informed consent will be implied. 
For participants only completing a telephone visit, assent/informed consent will be 
given verbally. 
4.2.2 Exclusion Criteria – LEAP Siblings 
There are no exclusion criteria for LEAP siblings. 
4.3 
LEAP PARENTS 
4.3.1 Inclusion Criteria – LEAP Parents 
Individuals must meet all of the following criteria to be eligible for this cohort in the 
study: 
1. Biological parent of LEAP participant.  
2. Willingness to participate in at least one study data collection (i.e. questionnaire, skin 
pricking testing, or blood draw). 

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3. Informed consent. For participants only returning a Questionnaire in any format, 
informed consent will be implied. For participants only completing a telephone visit, 
informed consent will be given verbally. 
4.3.2 Exclusion Criteria – LEAP Parents 
There are no exclusion criteria for LEAP parents. 
4.4 
PREMATURE TERMINATION OF A PARTICIPANT FROM THE STUDY 
Withdrawal of consent. Participants who withdraw consent for further procedures. 
These participants will be asked if they would be willing to complete an end-of-study 
visit to include specific assessments. 
Investigator decision. The principal investigator may choose to withdraw a participant 
from the study for any reason.  
Failure to return. Participants who do not return for visits and who do not respond to 
repeated attempts by the site staff to have them return will be considered lost to follow-
up.  
5. 
STUDY THERAPIES AND MEDICATIONS  
This section does not apply. 
6. 
STUDY ASSESSMENTS AND WINDOWS 
6.1 
VISIT WINDOWS 
6.1.1 Scheduled Visits 
The schedule of events is in Appendix 1. There is one visit, termed Visit 144. 
LEAP Participants 
Visit 144 must take place after the LEAP participant reaches 114 months (9.5 years) of 
age and before August 31, 2022. 
Siblings and Parents 
Visit 144 must take place before August 31, 2022. 
 
6.1.2 Window to Complete Visit 144 
More than one clinic appointment may be required to complete all assessments. The visit 
must be completed within 8 weeks of enrollment. The visit window does not apply to the 
telephone visit (Section 6.2.3). 
Participants may be asked to return to obtain additional samples in the event the samples 
were lost, damaged, or not sufficient. 
6.1.3 Definition of Enrollment and Study Participation 
Enrollment is the time at which a participant is determined to be eligible for participation 
and signs consent. Study participation is the period from signing consent to the 

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completion of Visit 144 for any single participant. Different members of a family may 
conclude their participation at different times. 
6.2 
VISIT TYPES 
6.2.1 Clinic 
Clinic visits will take place at the Clinical Trials Research Unit. Assessments listed in 
sections 6.3 to 6.7 will be conducted. 
6.2.2 Home 
Home visits will be undertaken when participants who are residents in the UK are unable 
to attend the Clinical Trials Research Unit. For these home visits the Guy’s and St. 
Thomas’ NHS Foundation Trust’s “Protection and Support of Lone Workers Procedure” 
will be in use. The informed consent process and assessments will be the same as those 
completed at a clinic appointment, as outlined in sections 6.3 to 6.7, except those listed 
below.  
• 
Peanut oral food challenges will only be performed at home for participants at low 
risk of a reaction. Any participant who qualifies as being at risk of a peanut reaction 
(as defined in section 6.7.1) will not have an oral food challenge at home.  
• 
The following assessments will not be conducted at home visits as equipment in the 
clinic is required to conduct the assessments: 
• 
TEWL 
• 
Spirometry 
• 
Anthropometry (skin fold thickness and waist circumference) 
6.2.3 Telephone 
Telephone visits will be undertaken when participants are not able to or do not wish to 
come into the clinic or have a home visit. The assessments will be the same as those 
completed at a clinic appointment, except as noted below. Informed consent will be given 
verbally during the telephone visit.  
Additionally, verbal consent will be requested for the clinic staff to obtain peanut SPT 
and IgE allergy assessment results performed in the last year by contacting the clinic 
where the tests were performed.  IgE results will be accepted for those undertaken in 
standardized labs using Thermo Fisher Scientific ImmunoCap System.The following 
assessments will not be conducted due to the visit not being conducted in person.  
• 
Physical Exam 
• 
SCORAD 
• 
Urine pregnancy test 
• 
TEWL 
• 
Spirometry 
• 
Allergy Assessments specified in Section 6.5, except those performed in the last 
year and obtained as specified above or obtained through home collection kits 
detailed below. 

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Participants who undergo a telephone visit may subsequently undergo an in person clinic 
visit or home visit. The assessments not included in the telephone visit will be conducted 
at the in person visit. In addition, parts of the telephone assessments may be repeated at 
the clinic visit or home visit at the discretion of the site investigator.  
For participants who are unable to attend the study site or undergo a home visit, blood 
may be collected at home to complete Allergy Assessments detailed in 6.5. Collection 
will be done via finger prick or home collection kit, using either a lancet finger prick 
device and/or micro-needle capillary blood collection device. The methodology for 
collection of the specimen and postal return to the Laboratory will be explained by 
telephone. Samples may be subsequently stored for analyses as detailed in section 7.  
 
6.3 
GENERAL ASSESSMENTS 
6.3.1 Non-dietary Assessments 
The following general assessments will be performed: 
• 
Informed consent: Informed consent will be obtained before any study procedures are 
performed. For children younger than 16, assent will also be obtained. 
• 
Eligibility criteria. 
• 
Medical history. 
• 
Physical examination: Temperature, pulse, respiratory rate, weight and height will be 
completed for LEAP participants and siblings <18 years of age. Skin fold thickness 
and waist circumference measurements will only be taken for LEAP participants. 
• 
Urine pregnancy test on all post-menarcheal participants prior to undergoing an 
incremental oral food challenge.  
• 
Concomitant medications: All concomitant medications will be recorded on the 
CRFs. 
• 
Eczema: SCORAD: Both subjective and objective eczema severity criteria will be 
recorded. The modified SCORAD evaluation alone will be used for all remaining 
eczema evaluations. 
• 
TEWL: this is a measure of water loss from skin and will be recorded for LEAP 
participants and siblings. 
• 
Bed dust collection: All families will be sent a vacuum pack with an information 
sheet, consent forms and instructions on how to collect a dust sample from bed-
sheets. 
• 
Rhinoconjunctivitis and Oral Allergy Syndrome (OAS) evaluation: Symptoms in 
accordance with the study definitions for seasonal and perennial rhinoconjunctivitis 
and OAS will be recorded. 
• 
Asthma: Symptoms in accordance with the study definition for asthma will be 
recorded and spirometry measurements taken for LEAP participants and siblings. 
• 
Food reaction history: A history will be taken to determine if the participant has had 
any clinically significant food-induced, immediate-onset allergic reactions. 

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• 
Food Allergy History Questionnaire: A questionnaire relating to the consumption of 
common food allergens (including tree nuts and sesame), foods that commonly cause 
OAS symptoms, and any other foods causing reactions indicative of immediate onset 
food allergy or OAS will be completed for all participants.  
• 
Adverse events: Participants will be assessed for adverse events. Adverse events will 
be recorded as described in section 8. 
6.3.2 Dietary Assessments 
• 
Dietary history: A dietary history will be obtained for LEAP participants only using a 
3-day food diary that captures typical food consumption around the time of the visit 
and provides a breakdown of macro- and micronutrient intake and total energy 
intake. 
• 
Peanut consumption monitoring: Peanut consumption questionnaires will be used to 
assess all participant’s peanut consumption. 
• 
Dietary education: Dietitians will provide dietary advice specific to each participant’s 
allergic profile. 
6.4 
CLINICAL LABORATORY ASSESSMENTS 
Routine clinical laboratory assessments are not planned in this study. 
6.5 
ALLERGY ASSESSMENTS 
• 
SPT for peanut (see section 6.6) 
• 
IgE total 
• 
IgE and IgE component resolved diagnosis (CRD) for peanut 
• 
IgG for peanut 
• 
IgG4 and IgG4 component resolved diagnosis (CRD) for peanut 
• 
Peanut oral food challenges  
• 
SPT to ingested allergens by extract to include hen’s egg white, tree nuts, sesame; 
fresh food to include hens egg white, and if milk and sesame equivocal on extract, 
fresh milk and tahini   
• 
SPT to inhaled allergens by extract to include Timothy grass, birch pollen, alternaria 
mould, cat and dog dander, and dust mite (Dermatophagoides pteronyssinus)  
• 
Sp-IgE to ingested allergens to include hen’s egg white, cow’s milk, peanut, sesame, 
Brazil nut, walnut, cashew and almond 
• 
Sp-IgE to inhaled allergens to include dust mite, cat, dog, alternaria, Timothy grass 
and birch pollen 
6.6 
SKIN-PRICK TESTING 
Because of the importance of skin prick testing in this study, some details regarding 
performing this testing are included here. 
Prior to testing, ensure that the participant has not received short-acting antihistamine 
medications for at least 48 hours and/or long-acting antihistamine medications for at least 
7 days. 

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The SPT for raw hen’s egg white will be performed using Red Lion salmonella-free egg. 
The other SPTs will be performed using Soluprick® extracts (ALK-Abelló, where 
available). Lyophilized peanut extract (ALK-Abelló) will be analyzed for total protein 
and major peanut allergen concentration (Ara h1, Ara h2, and Ara h3). The extract will be 
stored at −72 °C and will be used for SPT evaluations to peanut throughout the study. 
Saline 0.9% will be used as a negative control, and histamine (concentration of 1 mg/mL 
of saline) will be used as the positive control. 
Tests will be performed on the forearm unless unaffected eczema-free skin patches are 
not available, in which case the skin on the participant’s back will be used for testing. 
Using a standardized lancet (ALK-Abelló), the skin will be pricked through a drop of the 
extract, which will then be absorbed. 
Skin test sites should be measured after 15 minutes. The wheal and erythema should be 
measured at their widest diameters and recorded separately. Tests will be interpreted 
based on the widest wheal diameter. 
The positive and negative control tests should be performed and measured prior to 
allergen SPT. If the saline negative control test is ≥ 3 mm, then it should be repeated 
immediately. If the repeat test remains ≥ 3 mm, the testing should be rescheduled for 
approximately 7 days’ time. If the histamine positive control is ≤ 3 mm, then it should be 
repeated immediately. If the repeat test remains ≤ 3 mm, then the testing should be 
rescheduled for approximately 7 days’ time. 
For peanut measurements, the following rules apply: 
• 
The SPT will be performed in duplicate and the mean of the two tests will be 
recorded. 
• 
If both results are ≥ 1 mm and at least one result is ≤ 4mm, and there is a > 2 mm 
difference between the results, a third SPT will be performed and the mean of the two 
closest results will be recorded. 
• 
If one result is < 1 mm and one result is ≥ 1 mm, a third SPT will be performed. If 
two of three results are < 1 mm, 0 mm will be recorded as the final result. If two of 
three results are ≥ 1 mm, the mean of those two results will be recorded as the final 
result. 
• 
When both SPTs are >4mm, the average of the two peanut SPTs will be recorded 
without necessitating a third SPT even if the difference between the two SPTs is 
>2mm. If the first or second SPT is <4mm the existing protocol rules for a third SPT 
apply.  
6.7 
PEANUT CHALLENGES 
6.7.1 Conduct 
Peanut challenges will be conducted for LEAP participants and siblings <18 years of age 
as either open (unblinded) challenges or double-blind, placebo-controlled food challenges 
(DBPCFC). If a participant has a positive pregnancy test, the incremental oral food 
challenge will not be conducted. 

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Participants who are ≤ 36 months of age at the time of the challenge will consume 3.85 g 
peanut protein during the challenge. Participants who are > 36 months of age at the time 
of the challenge will consume 9.35 g peanut protein during the challenge. 
The open food challenges will be either cumulative (single dose) or incremental 
(sequentially increasing doses), depending on the participant’s risk of a peanut-induced 
allergic reaction. Participants are considered as being at risk of a peanut reaction if they 
meet one or more of the following criteria: 
• 
they have had a previous peanut-induced allergic reaction, 
• 
they have no known previous ingestion of peanut and also have a SPT (as per above 
methodology) of greater than 2 mm to peanut on the day of the challenge, or 
• 
the investigator suspects peanut allergy. 
 
If the participant is considered at risk of a peanut reaction (as defined above), the 
challenge will be performed as an incremental open challenge (see section 6.7.1.1.1). The 
participant will consume 3.85 g of peanut protein if they are ≤ 36 months of age or 9.35 g 
of peanut protein if they are > 36 months of age for the incremental open (unblinded) 
challenge. 
If the participant is considered at low risk of a peanut reaction, the challenge will be 
performed as a cumulative open (unblinded) challenge (see section 6.7.1.1.2). The 
participant will consume 3.85 g of peanut protein if they are ≤ 36 months of age or 9.35 g 
of peanut protein if they are > 36 months of age for the cumulative open (unblinded) 
challenge. The participant must consume 3.85 g of peanut protein if they are ≤ 36 months 
of age or a minimum of 5 g of peanut protein if they are > 36 months of age with no 
allergic symptoms to complete the cumulative open (unblinded) challenge. 
A DBPCFC will be performed when the open challenge results in an inconclusive 
outcome or at the discretion of the investigator (i.e. participant with food aversion or high 
anxiety). Participants > 36 months of age will consume a total of 9.35 g of peanut protein 
for the DBPCFC (4.35 g as blinded doses and 5 g as the final open dose). Participants ≤ 
36 months of age will consume a total of 3.85 g of peanut protein for the DBPCFC (1.85 
g as blinded doses and 2.0 g as the final open dose). 
A DBPCFC will preferentially be conducted on two separate days (see section 6.7.1.2.1). 
However, if the family can only attend for a single day, the DBPCFC will be undertaken 
as a single day mixed challenge with active and placebo doses randomly interspersed (see 
section 6.7.1.2.2). 
The peanut challenge will be discontinued if a reaction occurs that meets the study 
outcome criteria for a positive challenge (see Table 1 in section 6.7.2) and action will be 
taken according to local hospital guidelines. After discussion with an investigator, doses 
may be repeated if subjective or equivocal symptoms occur. 
6.7.1.1 Open challenges 
6.7.1.1.1 
Incremental open challenge 
For participants and eligible siblings (i.e. <18 years of age) considered at risk of a 
peanut reaction, as defined in section 6.7.1 above: 

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For participants ≤ 36 months of age (3.85 g): 
• 
Administer 3.85 g of peanut protein in five doses of 0.1, 0.25, 0.5, 1.0, and 2.0 g in 
separate meals. 
• 
For at risk participants, two lower doses of 0.01 g and 0.03 g peanut protein may be 
given prior to the usual 0.1 g starting dose at the discretion of the physician. 
• 
After each dose, observe the child for at least 15 minutes. If there is no reaction, 
administer additional peanut doses. 
 
For participants > 36 months of age (9.35 g): 
• 
Administer 9.35 g of peanut protein in six doses of 0.1, 0.25, 0.5, 1.0, 2.5, and 5.0 g 
in separate meals. 
• 
For at risk participants, two lower doses of 0.01 g and 0.03 g peanut protein may be 
given prior to the usual 0.1 g starting dose at the discretion of the physician. 
• 
After each dose, observe the child for at least 15 minutes. If there is no reaction, 
administer additional peanut doses. 
6.7.1.1.2 
Cumulative open challenge 
For participants and eligible siblings (i.e. <18 years of age) unlikely to have a peanut 
reaction : 
 
For participants ≤ 36 months of age (3.85 g): 
• 
Administer 3.85 g of peanut protein within a 6-hour period. The participant must 
consume complete dose with no allergic symptoms to complete the cumulative open 
(unblinded) challenge. 
• 
Observe the child during the challenge and for at least 1 hour after completion of the 
cumulative dose.  
 
For participants > 36 months of age (9.35 g): 
• 
Administer 9.35 g of peanut protein within a 6-hour period. The participant must 
consume a minimum of 5 g of peanut protein with no allergic symptoms to complete 
the cumulative open (unblinded) challenge. 
• 
Observe the child during the challenge and for at least 1 hour after completion of the 
cumulative dose.  
No particular feeding regimen is required. Peanut meals may vary and be used 
interchangeably. 
6.7.1.2 Double-blind, placebo-controlled food challenge 
DBPCFCs will be conducted for participants where their open (unblinded) challenge 
outcome is inconclusive, or at the investigator’s discretion. The total dose for participants 
≤ 36 months of age: 3.85g (1.85 g as blinded doses and 2.0g as the final open dose). The 

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total dose for participants > 36 months of age: 9.35 g (4.35 g as blinded doses and 5 g as 
the final open dose). 
Dose assessments and adjustments: 
o The active and placebo meals will be blinded by a computer-generated random 
code known to the dietician but not to the participant, nurse, or doctor. 
6.7.1.2.1 
Two-day separate double-blind, placebo-controlled food challenge 
Perform the two-day DBPCFC as follows: 
For participants ≤ 36 months of age (3.85 g): 
Day 1: 
o Administer four blinded doses in increasing increments of 0.1, 0.25. 0.5, and 1.0 
g, all of which are either peanut protein or placebo depending on the 
randomisation. Two lower doses of 0.01 g and 0.03 g peanut protein may be given 
prior to the 0.1 g dose at the discretion of the physician. 
o After each dose, observe the child for at least 15 minutes. 
Day 2: 
o Administer four blinded doses in increasing increments of 0.1, 0.25. 0.5, and 1.0 
g, all of which are either peanut protein or placebo depending on the 
randomisation. If peanut protein was administered on day 1, then administer 
placebo on day 2, or vice versa. A lower dose of 0.03 g peanut protein may be 
given prior to the 0.1 g dose at the discretion of the physician. 
o After each dose, observe the child for at least 15 minutes. 
o If the top blinded dose on day 2 is reached with no allergic reaction, wait 20 
minutes and administer an additional open (unblinded) 2.0 g dose of peanut 
protein. 
For participants > 36 months of age (9.35 g): 
Day 1: 
o Administer five blinded doses in increasing increments of 0.1, 0.25. 0.5, 1.0, and 
2.5 g, all of which are either peanut protein or placebo depending on the 
randomisation. Two lower doses of 0.01 g and 0.03 g peanut protein may be given 
prior to the 0.1 g dose at the discretion of the physician. 
o After each dose, observe the child for at least 15 minutes. 
Day 2: 
o Administer five blinded doses in increasing increments of 0.1, 0.25. 0.5, 1.0, and 
2.5 g, all of which are either peanut protein or placebo depending on the 
randomisation. If peanut protein was administered on day 1, then administer 
placebo on day 2, or vice versa. A lower dose of 0.03 g peanut protein may be 
given prior to the 0.1 g dose at the discretion of the physician. 

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o After each dose, observe the child for at least 15 minutes. 
o If the top blinded dose on day 2 is reached with no allergic reaction, wait 20 
minutes and administer an additional open (unblinded) 5 g dose of peanut protein. 
6.7.1.2.2 
One-day mixed double-blind, placebo-controlled food challenge 
Perform the one-day DBPCFC as follows: 
For participants ≤ 36 months of age (3.85 g): 
o Administer at least 1.85 g of peanut protein in four blinded doses, randomly 
interspersed with 1 placebo dose of equivalent portion size to the previous peanut 
dose, in increasing increments of 0.1, 0.25, 0.5, and 1.0 g (i.e. 5 separate meals of 
either peanut or placebo will be given over the course of 1 day). Two lower doses 
of 0.01 g and 0.03 g peanut protein may be given prior to the 0.1 g dose at the 
discretion of the physician. 
o After each dose, observe the child for at least 15 minutes.  
o If the top blinded dose is reached with no allergic reaction, wait at least 20 
minutes and administer an additional open (unblinded) 2.0 g dose of peanut 
protein. 
o An additional dose pair comprising a repeat of the previous dose and a placebo in 
random order may be given at the discretion of the investigator. 
For participants > 36 months of age (9.35 g): 
o Administer at least 4.35 g of peanut protein in five blinded doses, randomly 
interspersed with 3 placebo doses of equivalent portion size to the previous peanut 
dose, in increasing increments of 0.1, 0.25, 0.5, 1.0, and 2.5 g (i.e. 8 separate 
meals of either peanut or placebo will be given over the course of 1 day). Two 
lower doses of 0.01 g and 0.03 g peanut protein may be given prior to the 0.1 g 
dose at the discretion of the physician. 
o After each dose, observe the child for at least 15 minutes.  
o If the top blinded dose is reached with no allergic reaction, wait at least 20 
minutes and administer an additional open (unblinded) 5 g dose of peanut protein. 
o An additional dose pair comprising a repeat of the previous dose and a placebo in 
random order may be given at the discretion of the investigator. 
6.7.2 Outcome 
Outcome of the challenge will be determined by evaluating the participant using the 
criteria in Table 1. 

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Table 1. Criteria for determining the outcome of food challenge 
Major Criteria 
Confluent erythematous pruritic rash 
Respiratory signs (at least one of the following): 
Wheezing 
Inability to speak 
Stridor 
Dysphonia 
Aphonia 
≥3 Urticarial lesions 
≥1 Site of angioedema 
Hypotension for age not associated with vasovagal episode 
Evidence of severe abdominal pain (such as abnormal stillness or doubling over) that persists for ≥3 
minutes 
Minor Criteria 
Vomiting 
Diarrhea 
Persistent rubbing of nose or eyes that lasts for ≥ 3 minutes 
Persistent rhinorrhea that lasts for ≥ 3 minutes 
Persistent scratching that lasts for ≥ 3 minutes 
 
A positive food challenge will be defined by the presence of either of the following: 
• One or more major criteria. 
• Two or more minor criteria. 
An indeterminate food challenge will be defined by the presence of only one minor 
criterion. In this case the oral food challenge may repeated on a separate occasion. 
A negative food challenge will be defined by the absence of major or minor criteria. 
All symptoms should be of new onset and not due to ongoing disease. Symptoms must 
occur no later than 2 hours after the last dose. 
7. 
TOLERANCE ASSAYS 
7.1 
MECHANISTIC HYPOTHESES 
Several hypotheses related to potential mechanisms of peanut allergy will be explored in 
this trial. There is good evidence that peanut allergy is characterized by Th2-skewing and 
production of IgE to peanut proteins13.  
Peanut-specific CD4+ T cells are known to be involved in the pathophysiology of peanut 
allergy.14,13 New reagents that allow quantification and phenotyping of these cells using 
Ara h1-specific class II tetramers15 with standard flow cytometry markers will allow us to 
monitor frequencies and phenotypes of peanut-reactive T cells in relation to peanut 
allergy status. 
A report by Wambre et al.16 suggests that a unique subset of Th2 cells may be a 
biomarker for allergy, and that a decrease in this cell subset may be indicative of 
desensitization or tolerance to peanut. We intend to monitor the frequency of these cells, 

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denoted Th2A, to determine if modification of this cell subset could serve as a surrogate 
biomarker for peanut allergy. 
Based on these findings the mechanistic assays outlined in subsequent sections are 
proposed. 
7.2 
PROPOSED MECHANISTIC ASSAYS 
7.2.1 Serum/Plasma Assays 
Immunoglobulin assays: IgE, IgG, and IgG4 anti-peanut will be measured on an 
ImmunoCAP™ instrument (Phadia) or equivalent over time in subjects followed from 
the primary LEAP trial and between groups. 
Component-resolved assays: Phadia’s ImmunoCAP™ peanut component tests for 
quantification of IgE and IgG4 against specific peanut allergens, such as Ara h1, 2, 3, 6, 
and 8 and may be used to evaluate changes in specific reactivity over time in subjects 
followed from the original LEAP trial and between groups. 
Epitope arrays: Epitope-specific IgE and IgG4 in the plasma may be measured for Ara 
h1, 2, and 3, as previously described.17 The arrays include 20-mer peptides offset by three 
amino acids, and cover the entire sequence of these three major peanut allergens. Note 
that this approach does not measure conformational epitopes.  
Note that for direct quantitative comparison with LEAP and LEAP-On data, it is optimal 
to retest plasma from all visits for the same subjects in the previous trials together with 
the current trial pending sample availability. 
IgE-Facilitated antigen binding (FAB) assay: The IgE-FAB inhibition assay uses 
fluorescence-activated cell sorting to measure serum inhibitory activity for IgE-facilitated 
CD23-dependent binding of allergen-IgE complexes to B cells (IgE-FAB), possibly due 
to blocking antibodies.18 Vickery et al. have shown reduced IgE-FAB in patients treated 
with peanut oral immunotherapy.19 We hypothesize that persistent increases in serum 
inhibitory activity for IgE-FAB correlate with desensitization and tolerance. Longitudinal 
plasma samples from the LEAP participants at LEAP Visit 60 and visit 144 in the current 
study, and from their siblings may be assessed for inhibitory activity on the IgE-FAB 
assay as a functional measure of blocking antibodies in plasma potentially induced by 
peanut consumption.  
IgE-Mast cell activation test (MAT) /inhibition (iMAT) assays:  Dr. Alexandra Santos 
has developed a novel mast cell degranulation assay using the LAD2 mast cell line where 
cells are sensitized with plasma from patients and the expression of activation markers 
following stimulation with allergen is assessed by flow cytometry: the mast cell 
activation test (MAT). This experimental system reflects in vitro the clinical reactivity or 
tolerance to peanut20 and is currently being validated in a larger (n=165) existing set of 
samples of patients previously assessed for peanut allergy. The inhibition of mast cell 
activation test (iMAT) assesses the ability of plasma to interfere with peanut-induced 
degranulation of mast cells previously sensitized with plasma from a peanut allergic 
patient.  

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Santos AF et al.20 demonstrated that plasma from peanut-sensitized but tolerant patients 
and plasma from peanut-allergic patients who underwent peanut oral immunotherapy was 
able to reduce peanut-induced activation of mast cells that had been previously sensitized 
with plasma from peanut allergic patients. We hypothesize that inhibition of peanut-
induced mast cell activation, possibly due to blocking antibodies, is associated with 
desensitization and tolerance. MAT and iMAT could be performed by the laboratory of 
Dr. Alexandra Santos. Plasma samples collected at Visit 144 from LEAP participants and 
their siblings could be tested by MAT together with samples collected at LEAP visit 60 
to assess the ability of IgE antibodies present in the plasma samples to induce mast cell 
activation following stimulation with peanut extract. The same samples can be tested by 
iMAT to assess the ability of blocking antibodies to inhibit peanut-induced mast cell 
activation. The results of MAT and iMAT will be interpreted in relation to peanut allergy 
and tolerance.  
7.2.2 Cellular Assays 
Blood will be collected for PBMC isolation as specified in the SOE and sent to the ITN 
designated core laboratory for processing using ITN approved standard operating 
procedures (SOPs) for PBMC separation and aliquoting. This ensures that standardized 
procedures are used and that high quality material is obtained for testing. PBMCs will be 
stored in the vapor phase of liquid nitrogen until use. These cells may be used as 
described below and will be available for future studies such as immunosequencing and 
functional studies when remaining cells’ numbers are sufficient to make these assays 
technically feasible.  
Tetramer assays: We anticipate that tetramer assays will be done in collaboration with 
Dr. W. Kwok at the Benaroya Research Institute in Seattle, WA, USA. Dr. Kwok has 
generated and tested tetramer reagents for Ara h1, 2, 3, 6, and 8. For example, tetramers 
for Ara h1, restricted by eight HLA class II alleles, have been successfully used to 
examine the frequency and phenotype of peanut-specific CD4+ T cells in individuals 
with and without peanut allergy.15 These reagents have been successfully used to stain 
previously frozen PBMCs. The ability to use these reagents will depend on the overlap 
between HLA alleles among study participants and available reagents. These and similar 
reagents can be used to track changes in frequency of CD4+ T cells in response to 
therapy. For optimum use, these assays currently require 20 million viable cells per assay, 
so it is important that every effort be made to collect the full planned blood volumes at 
the time points specified the SOE. 
Since tetramers are not currently available for all haplotypes, other T-cell assays may also 
be performed such as the CD154 up-regulation assay for monitoring peanut allergen-
reactive CD4 T cells with Erik Wambre at Benaroya Research Institute in Seattle, WA, 
USA. This assay currently requires overnight in vitro stimulation of 20-40 million 
PBMCs with peanut allergen extract in the presence of anti-human CD40 blocking mAb. 
Half of the cells can then be collected and analyzed by flow cytometry, and the other half 
can be used for sorting CD154+ and CD154- CD4 T cells for RNAseq analysis.  
T cell immunoprofiling: T cell immunoprofiling may be carried out to determine if 
parallel mechanisms of anergy, exhaustion, deviation, induced regulation, or deletion are 
occurring in the peanut-specific T cell subset. Live/dead (for deletion evaluation) and 

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apoptosis marker staining can occur in parallel with phenotyping and sorting of tetramer-
positive or CD154-positive CD4 T cells for RNAseq analysis.  
TH2A subset analysis: We may also use surface flow cytometry to determine the 
frequency of TH2A cells15 as this subset of Th2 cells may be a biomarker for allergy. 
This assay has the advantage that it can be reliably performed with only one million 
previously frozen, viable PBMCs. Work by Wambre16 suggests that this subset includes 
the vast majority of allergen-specific CD4+ T cells as determined by tetramer analysis. 
Other immune cells: Through high throughput multiplex immunoprofiling, we will be 
able to determine absolute counts of subsets of other immune cells such as dendritic cells, 
natural killer T cells, and others. 
7.2.3 DNA Assays 
DNA-HLA genotypes: MHC tetramers bind to the T-cell receptor in an HLA-specific 
context. Therefore, DNA may be isolated from participants’ peripheral blood cells to 
perform sequence-based HLA typing, so that appropriate candidates may be identified for 
tetramer analysis as described in section 7.2.2. Tetramer reagents for peanut have been 
focused on the HLA class II molecules DRB1 and DRB3. However, we may type DQ 
and DP as well as DR alleles in the event that new data suggests that those alleles are also 
important. 
7.2.4 Gene Expression Profiling 
Gene expression profiling may be performed on RNA isolated from peripheral whole 
blood using RNAseq, nanostring, high-throughput real-time PCR, or other methods.  The 
goal of these assays would be to identify differences in transcriptomic profiles and assess 
relatedness to cytokine levels, cellular profiles, or other study characteristics.   
7.3 
FUTURE AND UNPLANNED STUDIES 
Retention of Samples  
A major priority of the Immune Tolerance Network, in partnership with the National 
Institute of Allergy and Infectious Diseases of the NIH, USA, is the development of 
novel immunoassays in order to better understand mechanisms of tolerance and to 
develop biomarkers to predict the development and maintenance of clinical tolerance. As 
in all Immune Tolerance Network-supported clinical trials, informed consent will be 
obtained from all participants for their samples to be stored for use in future studies.  
Biological specimens collected in this trial will be stored long-term in order to re-
evaluate biologic responses as new research tools to study tolerance become available. 
The blood specimens will therefore be stored at the ITN sample repositories located in 
Indianapolis, IN, USA and Piscataway, NJ, USA for a minimum of 10 years.  
Residual specimens may be used by the investigators for development of new 
immunologic assays or for cross-trial comparisons. Although specimens in this protocol 
are described in the context of assays to be performed, it should be noted that not 
necessarily all assays will be performed for all participants at each time point. Decisions 
to perform assays will be made based on statistical and scientific planning, hypotheses to 
be tested, and technologies available. Finally, clinical outcomes will be taken into 

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account to determine the potential value of the assays. For example, if a clinical effect 
fails to occur, it may be decided that there is minimal value in performing certain 
mechanistic assays.  
8. 
ADVERSE EVENTS 
8.1 
OVERVIEW 
This section defines types of Adverse Events (AEs) and outlines the procedures for 
appropriately collecting, grading, recording, and reporting them. Information in this 
section complies with ICH Guideline E2A: Clinical Safety Data Management: 
Definitions and Standards for Expedited Reporting; and ICH Guideline E-6: Guidelines 
for Good Clinical Practice.  
This is a trial with a single planned study visit. This visit takes place during a window 
between enrollment and completion of the visit (section 6.1.2). As a cross-sectional 
study, it does not aim to capture disease or therapy-related adverse events over time. 
Medical history and other study tools will provide retrospective information related to 
allergy and related conditions. 
Adverse events will however be collected during the period delineated in section 8.3.1 
Collection Period. The principal aim is to capture adverse events that may be related to 
study procedures such as venipuncture, home self blood collection, skin-prick testing, 
assessment of TEWL, and oral food challenges. 
With these parameters, the study investigator is responsible for the detection and 
documentation of events meeting the definition of an AE (adverse event) or SAE (serious 
adverse event) as described in sections 8.2. All of these procedure-related AEs will be 
recorded in the source documents and on electronic CRF(s). All serious adverse events 
(SAEs) will be reported on a Serious Adverse Event Report Form as well as on electronic 
CRFs. In addition, SAEs will be reported in accordance with local investigative site 
guidelines. 
Adverse events that are classified as serious according to the definition of health 
authorities must be reported within 24 hours of learning of the event to the NIAID and 
the ITN. 
8.2 
DEFINITIONS 
8.2.1 Adverse Event 
An adverse event is any occurrence or worsening of an undesirable or unintended 
sign, symptom, laboratory finding, or disease that occurs during participation in 
the study. An adverse event that occurs during the study visit will be followed until it 
resolves or until 30 days after a participant terminates from the study, whichever comes 
first.  
8.2.2 Serious Adverse Event 
An AE is considered “serious” if, in the view of either the investigator or DAIT/NIAID it 
results in any of the following outcomes (21 CFR 312.32(a)): 
1. Death  

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2. A life-threatening event: An AE is considered “life-threatening” if, in the view of 
either the investigator or DAIT/NIAID, its occurrence places the subject at 
immediate risk of death. It does not include an AE that, had it occurred in a more 
severe form, might have caused death.  
3. Inpatient hospitalization or prolongation of existing hospitalization. 
4. Persistent or significant incapacity or substantial disruption of the ability to conduct 
normal life functions. 
5. Congenital anomaly or birth defect. 
6. Important medical events that may not result in death, be life threatening, or require 
hospitalization may be considered serious when, based upon appropriate medical 
judgment, they may jeopardize the subject and may require medical or surgical 
intervention to prevent one of the outcomes listed above. 
8.3 
COLLECTING AND RECORDING ADVERSE EVENTS 
8.3.1 Collection Period  
Adverse events, as defined in this protocol, will be collected from the time the participant 
signs the informed consent until the participant completes study participation.  
Information regarding individual adverse events will be collected until the event is 
resolved or until 30 days after the participant completes study participation, whichever 
comes first.  
8.3.2 Collecting Adverse Events 
Adverse events may be discovered through any of these methods: 
• 
Observing the participant. 
• 
Questioning the participant in an objective manner. 
• 
Receiving an unsolicited complaint from the participant. 
In addition, an abnormal value or result from a clinical or laboratory evaluation can also 
indicate an adverse event. If an abnormal value or result is determined by the investigator 
to be clinically significant, it must be recorded as an adverse event on the appropriate 
laboratory evaluation form. 
8.3.3 Recording Adverse Events 
Throughout the study, the investigator will record all AEs, as defined by the protocol, on 
the appropriate electronic CRF. The investigator will treat participants experiencing AEs 
appropriately and observe them at suitable intervals until their symptoms resolve or their 
status stabilizes. 
8.3.4 Recording Serious Adverse Events 
Serious adverse events related to any procedure will be recorded on the adverse event 
source document form and e-CRF. 

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8.4 
GRADING AND ATTRIBUTION OF ADVERSE EVENTS 
8.4.1 Grading Criteria 
The study site will grade the severity of AEs experienced by study participants according 
to the criteria set forth in the National Cancer Institute‘s Common Terminology Criteria 
for Adverse Events 4.03 (published June 14, 2010). This manual provides a common 
language to describe levels of severity, to analyze and interpret data, and to articulate the 
clinical significance of all AEs. 
Adverse events will be graded on a scale from 1 to 5 according to the following standards 
in the CTCAE manual: 
• Grade 1 = mild  
• Grade 2 = moderate  
• Grade 3 = severe  
• Grade 4 = life-threatening  
• Grade 5 = death 
For additional information and a printable version of the CTCAE manual, go to 
http://ctep.cancer.gov/reporting/ctc.html.  
8.4.2 Attribution  
The site investigator will make the initial determination of the relation, or attribution, of 
an AE to study procedures and will record the initial determination on the appropriate 
CRF and/or SAE reporting form. The relation of an AE to study procedures will be 
determined using definitions in Table 2. Final determination of attribution for safety 
reporting will be decided by DAIT/NIAID.  
Table 2. Attribution of Adverse Events 
Code 
Descriptor 
Relationship to study procedure 
Unrelated Category 
1 
Not Related 
The adverse event is clearly not related: there is insufficient 
evidence to suggest a causal relationship. 
Related Categories 
2 
Possible 
The adverse event has a reasonable possibility to be related; 
there is evidence to suggest a causal relationship. 
3 
Related 
The adverse event is clearly related. 
 
8.5 
REPORTING SERIOUS ADVERSE EVENTS 
8.5.1 Reporting SAEs to the Sponsor  
The following process for reporting an SAE ensures compliance with 21CFR 312 and 
ICH guidelines. After learning that a participant has experienced an SAE, the investigator 
or designee will report the SAE to the DAIT/NIAID Statistical and Clinical Coordinating 
Center (SACCC, Rho Federal) via the electronic SAE report form (SAE CRF) within 24 
hours of becoming aware of the event. The initial SAE CRF should include as much 
information as possible, but at a minimum must include the following: 

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• AE term 
• Study procedure 
• Reason why the event is serious 
• Supplementary CRF pages that must be current at the time of SAE reporting: medical 
history, concomitant medications, demographics, death.  
As additional details become available, the SAE CRF should be updated and re-
submitted. Every time the SAE CRF is submitted, it should be electronically signed by 
the study investigator or sub investigator. 
For additional information regarding SAE reporting, contact Rho Product Safety: 
Rho Product Safety  
2635 East NC Highway 54 
Durham, NC  27713 
Toll-free - (888) 746-7231 
SAE Fax Line: 1-888-746-3293 
Email: rho_productsafety@rhoworld.com 
8.5.2 Reporting SAEs to the DSMB 
The DAIT/NIAID will provide the NIAID DSMB with information regarding all SAEs 
on a regular basis as determined by the DAIT/NIAID Medical Monitor. SAE’s will also 
be reviewed during the regular annual NIAID DSMB safety reviews described in section 
3.3.3. 
8.5.3 Reporting SAEs and Unanticipated Problems to the Ethics Committee 
The investigators must report SAEs and unanticipated problems to their local Ethics 
Committee according to local guidelines.  
9. 
STATISTICAL CONSIDERATIONS AND ANALYTICAL PLAN 
9.1 
ANALYSIS SAMPLES 
The analysis samples are defined separately for each cohort. The three cohorts are LEAP 
participants, siblings of LEAP participants, and parents of LEAP participants. 
LEAP Participants 
Intent to treat (ITT) sample:  any participant who enrolled in the LEAP Participant 
cohort who is evaluable for allergy. 
Per protocol (PP) sample:  any participant in the ITT sample who was in the LEAP PP 
sample. 
Safety sample (SS):  There is no safety sample for this cohort. 
 
LEAP Siblings 
Intent to treat (ITT) sample: any participant who enrolled in the LEAP Sibling cohort 
for whom at least one assessment is available. 
Per protocol (PP) samples:  any participant in the ITT sample whose sibling was in the 
LEAP PP sample. 

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Safety sample (SS):  There is no safety sample for this cohort. 
 
LEAP Parents 
Intent to treat (ITT) sample: any participant who enrolled in the LEAP parent cohort 
for whom at least one assessment is available. 
Per protocol (PP) samples:  There is no per-protocol sample for this cohort. 
Safety sample (SS):  There is no safety sample for this cohort. 
9.2 
ANALYSIS OF ENDPOINTS 
9.2.1 Primary Endpoint 
LEAP Participants 
Between-group comparison 
The primary endpoint is the rate of peanut allergy at 144 months of age (V144). The main 
comparison is of the rate between LEAP consumers and LEAP avoiders at V144. This 
comparison will be made by an unadjusted logistic regression model at the 0.05 level of 
significance. This comparison evaluates whether the consumption of peanut starting in 
infancy induces long-lasting tolerance to peanut. 
Analysis of transient desensitisation 
The within group comparison in LEAP Consumers between V72 and V144 will allow 
identification and statistical evaluation of a group of children with transient 
desensitisation who develop peanut allergy after ad libitum consumption. This matched 
pre-post test allows us to explore mechanistic and immunologic differences between 
those who appear to be transiently desensitised versus those who appear to be persistently 
tolerant. If there is a statistically significant increase in the rate of peanut allergy from 
V72 to V144, this will be interpreted as evidence of 'transient desensitisation'. This 
comparison of the proportion of participants with peanut allergy at V72 and V144 within 
the LEAP Consumption group will be made using a paired (pre/post) McNemar's test at a 
0.05 level of significance. 
The main analyses will be in the ITT population. 
Additional analyses will be performed to adjust for confounders, such as age, gender and 
others. 
LEAP Siblings 
The primary endpoint is the rate of peanut sensitization in younger siblings who resided 
in the home of the LEAP participant on or before LEAP Visit 60, as assessed in the 
current LEAP Trio study. The main comparison is of the sensitization rate between the 
younger siblings of LEAP consumers and LEAP avoiders. 
This comparison will be made by a logistic regression model with a random family effect 
included to account for the clustering of siblings within family units and the analysis of 
multiple younger siblings. This comparison evaluates whether the younger siblings of the 
LEAP consumers are at higher risk of peanut sensitization compared to the younger 

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siblings of LEAP avoiders due to the environmental exposure to peanut in conjunction 
with a lack of peanut consumption.  
Additional analyses will be done in older siblings. 
9.2.2 Secondary and Exploratory Endpoints 
Secondary 
Endpoint 
LEAP Participant Analysis 
LEAP Sibling Analysis 
LEAP Parent Analysis 
Clinical 
1,2,3,4* 
The amount of peanut 
consumption, SCORAD, skin-
prick test wheal sizes, and 
specific-IgE measurements 
including IgE to Ara h2 and 
other components will be 
compared between LEAP 
avoiders and LEAP consumers 
using parametric and non-
parametric tests (e.g. Wilcoxon 
and t-test). The data will first be 
tested to determine normality. If 
the data are not normal then a 
Wilcoxon test will be run. If the 
data are normal then tests will 
be run to determine if the 
variances are equal between the 
two groups. If the variances are 
equal, a two sample t-test 
assuming equal variances will 
be used; otherwise, a two 
sample t-test assuming unequal 
variances will be used. 
 
The amount of peanut 
consumption, SCORAD, skin-
prick test wheal sizes, and 
specific-IgE measurements 
including IgE to Ara h2 and 
other components will be 
compared between the siblings 
of the LEAP avoiders and LEAP 
consumers using linear mixed 
effects models with a random 
family effect included to 
account for the clustering of 
siblings within family units and 
the analysis of multiple siblings. 
The amount of peanut 
consumption, skin-prick test 
wheal sizes, and specific-IgE 
measurements will be compared 
between the parents of the 
LEAP avoiders and LEAP 
consumers using linear mixed 
effects models with a random 
family effect included to 
account for the clustering of 
parents within family units. 
5,6+ 
The prevalence of asthma, 
rhinitis and eczema will be 
compared between LEAP 
avoiders and LEAP consumers 
using a two-tailed, chi-squared 
test at the 0.05 level of 
significance. 
The prevalence of asthma, 
rhinitis, and eczema will be 
compared between the siblings 
of the LEAP avoiders and LEAP 
consumers using a logistic 
regression model with a random 
family effect included to 
account for the clustering of 
siblings within family units and 
the analysis of multiple siblings. 
 
The prevalence of asthma, 
rhinitis, and eczema will be 
compared between the parents 
of the LEAP avoiders and LEAP 
consumers using a logistic 
regression model with a random 
family effect included to 
account for the clustering of 
parents within family units. 
7 
 
The proportion of participants 
with peanut allergy will be 
compared between the siblings 
of the LEAP avoiders and LEAP 
consumers using a logistic 
regression model with a random 
family effect included to 
account for the clustering of 
siblings within family units and 
the analysis of multiple siblings. 
 
The proportion of participants 
with peanut allergy will be 
compared between the parents 
of the LEAP avoiders and LEAP 
consumers using a logistic 
regression model with a random 
family effect included to 
account for the clustering of 
parents within family units. 
 
8 
The proportion of participants 
experiencing at least one 
The proportion of participants 
experiencing at least one 
 

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Secondary 
Endpoint 
LEAP Participant Analysis 
LEAP Sibling Analysis 
LEAP Parent Analysis 
peanut-related adverse event 
will be compared between 
LEAP avoiders and LEAP 
consumers using a logistic 
regression model. 
 
peanut-related adverse event 
will be compared between the 
siblings of the LEAP avoiders 
and LEAP consumers using a 
logistic regression model with a 
random family effect included 
to account for the clustering of 
siblings within family units and 
the analysis of multiple siblings. 
 
Exploratory 
1 
 
The proportion of participants 
with food allergies other than 
peanut will be summarized 
descriptively.  
 
 
The proportion of siblings with 
food allergies other than peanut 
will be summarized 
descriptively.  
 
 
The proportion of parents with 
food allergies other than peanut 
will be summarized 
descriptively.  
 
2 
Dietary assessments will be 
compared between LEAP 
avoiders and LEAP consumers 
using parametric and non-
parametric tests (e.g. Wilcoxon 
and t-test). The data will first be 
tested to determine normality. If 
the data are not normal then a 
Wilcoxon test will be run. If the 
data are normal then tests will 
be run to determine if the 
variances are equal between the 
two groups. If the variances are 
equal, a two sample t-test 
assuming equal variances will 
be used; otherwise, a two 
sample t-test assuming unequal 
variances will be used. 
 
 
3 
TEWL will be compared 
between LEAP avoiders and 
LEAP consumers using 
parametric and non-parametric 
tests (e.g. Wilcoxon and t-test). 
The data will first be tested to 
determine normality. If the data 
are not normal then a Wilcoxon 
test will be run. If the data are 
normal then tests will be run to 
determine if the variances are 
equal between the two groups. If 
the variances are equal, a two 
sample t-test assuming equal 
variances will be used; 
otherwise, a two sample t-test 
assuming unequal variances will 
be used. 
 
TEWL will be compared 
between the siblings of the 
LEAP avoiders and LEAP 
consumers using linear mixed 
effects models with a random 
family effect included to 
account for the nesting of 
siblings within family units and 
the analysis of multiple siblings. 
 
*Secondary clinical endpoint #4 is only collected in LEAP participants and siblings of LEAP 
participants 

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+Secondary clinical endpoint #6 is only collected in the parents of LEAP participants 
 
Exploratory endpoint #4 (household peanut consumption and concentration of peanut 
dust) will be compared between the families of the LEAP avoiders and LEAP consumers 
using linear mixed effects models with a random family effect included to account for the 
nesting of multiple family members within family units. 
Younger and older siblings of LEAP participants will be analyzed separately. 
9.2.3 Safety Analysis 
Safety will be analyzed through the reporting and analysis of AEs. All AEs will be 
classified by body system and preferred term according to a standardized thesaurus. The 
severity of AEs will be classified using the NCI-CTCAE toxicity scale. The total number 
of events and the number of participants experiencing AEs will be summarized by body 
system and preferred term for each group and overall. Adverse events will also be 
summarized by maximum severity, relation to underlying disease, and relation to the 
study procedure for each group and overall. Separate summaries will be provided for 
serious AEs, procedure-related AEs, and AEs leading to study discontinuation. 
9.2.4 Relevant Medical History 
Relevant medical history within the past 12 months, including the existence of current 
signs and symptoms, will be collected for each body system. 
9.2.5 Use of Medications 
All medications taken by or administered to study participants beginning 30 days before 
enrollment and continuing throughout the study will be collected. All medications used 
will be coded according to the WHO drug dictionary. The number and percentage of 
participants receiving prior and concomitant medications/therapies will be presented 
overall and by medication class. 
9.3 
SAMPLE SIZE  
LEAP Participants 
The sample size is based on the number of children likely to be available to enroll in the 
study. As shown in the Figure 1 LEAP Screening, Participant and Sibling Cohorts, there 
are 311 LEAP avoiders and 319 LEAP consumers (total 630) potentially eligible for 
enrollment. In the LEAP study, the rate of peanut allergy was 3.2% among consumers 
and 17.2% among avoiders at 60 months of age. We assume that the rates will not change 
markedly between month 60 and 144. We wish to detect a difference between the groups 
that are similar to that detected at 60 months.3 88.5% of those potentially eligible enrolled 
in the follow on LEAP-On Study.5 We assume a lower percentage of those potentially 
eligible will enroll in this study. 
For example, if 80% of those potentially eligible enroll, the true rate of allergy among 
LEAP consumers at 144 months is 6%, and the true rate of allergy among LEAP avoiders 
at 144 months is 15%, there is 91% power to detect a difference with a two-sided type I 
error of 0.05 between the groups. If only 70% enroll and other assumptions remain 
constant, there would be about 87% power to detect a difference. 

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Other examples are shown in Figure 3. 
 
Figure 3. Power estimates for allergy rates 
Three possible recruitment rates are shown in the left, middle and right panels. In each panel, 
the power to detect a difference, with a two-sided type I error of 0.05, between groups is shown 
on the vertical axis for different assumptions regarding the true underlying allergy rates in the 
consumer and avoider groups at month 144. In each panel contour lines display rates in 
consumers of 2%, 4%, 6%, 8% and 10%  while the horizontal axis displays a range of rates in 
avoiders of 10 to 25%  
 
LEAP Siblings 
The sample size for LEAP siblings is based on observing as many individuals as is 
practical among those who can meet the eligibility criteria. Different sample sizes 
provide power to detect differences of varying amounts depending on how many 
individuals are available for analysis. 
The primary endpoint for the sibling cohort is listed in section 3.2. 
For protocol versions 1.0 and 2.0, we estimated that 222 younger siblings with at least 
one year of exposure to the LEAP intervention are potentially available from the LEAP 
avoidance arm and 207 from the LEAP consumption arm. In protocol version 3.0, we 
removed the criterion that younger siblings needed to have at least one year of exposure 
to the LEAP intervention. This resulted in an estimated 80 additional younger siblings to 
be available for recruitment into LEAP Trio. 

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We aim to be able to detect an increase in the sensitization rate of individuals meeting the 
primary endpoint between 18.4% from the siblings in the consumption arm and 8.4% 
from the siblings in the LEAP avoidance arm.  
These assumptions are derived from findings in a survey questionnaire regarding allergy 
among siblings: 9.2% among consumer siblings and 4.2%% among avoider siblings. The 
conversion rate from peanut allergy to sensitization [IgE ≥ 0.35, SPT ≥3mm] within the 
LEAP avoidance group at 60 months was 1.95 fold. At 72 months, the conversion rate 
within the LEAP avoidance group was 2.05 fold. The conversion rate of two fold (an 
average of the 60 and 72 month rates) was chosen to estimate the sensitization rate in 
younger siblings. For example, if 90% of the available individuals are enrolled and 
assessed, the rate of subjects meeting the primary endpoint [IgE ≥ 0.35, SPT ≥ 3mm] is 
8% in the siblings of LEAP avoiders, and the rate of those meeting the primary endpoint 
[IgE ≥ 0.35, SPT ≥ 3mm] is 17% in the siblings of LEAP consumers, there is 83% power 
to detect a difference with a two-sided type I error rate of 0.05 between the groups. If 
only 80% of the available individuals are enrolled and assessed and other assumption 
remain constant, there would be about 79% power to detect a difference and only 73% 
power if 70% of the available individuals are enrolled and assessed while the other 
assumptions remain constant. 
Other examples are shown in Figure 4. 

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Figure 4. Power estimates for sensitization rates  
Three possible recruitment rates for younger siblings are shown in the left, middle and right 
panels. In each panel, the power to detect a difference, with a two-sided type I error of 0.05, 
between groups is shown on the vertical axis for different assumptions regarding the true 
underlying rates for the primary endpoint among consumer and avoider younger siblings. In 
each panel contour lines display assumed rates in avoiders’ siblings of 4%, 6%, 8%, 10% and 
12% while the horizontal axis displays a range of rates in consumers’ sibling from 15% to 25%. 
 
9.4 
MISSING DATA 
Detailed methods for addressing missing data will be specified in the SAP. 
9.5 
REPORTING DEVIATIONS FROM THE ORIGINAL STATISTICAL PLAN 
The principal features of both the study design and the plan for statistical data analysis 
are outlined in this protocol and in the SAP. Any change in these features requires either 
a protocol or an SAP amendment, which is subject to review by the NIAID DSMB, the 
study sponsor and the Ethics Committee. These changes will be described in the final 
study report as appropriate. 
10. 
ACCESS TO SOURCE DATA/DOCUMENTS 
The investigational sites participating in this study will maintain the highest degree of 
confidentiality permitted for the clinical and research information obtained from 
participants in this clinical trial. Medical and research records should be maintained at 

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each site in the strictest confidence. However, as a part of the quality assurance and legal 
responsibilities of an investigation, the investigational sites must permit authorized 
representatives of the sponsors to examine (and to copy when required by applicable law) 
clinical records for the purposes of quality assurance reviews, audits, and evaluation of 
the study safety and progress. Unless required by the laws permitting copying of records, 
only the coded identity associated with documents or other participant data may be 
copied (and any personally identifying information must be obscured). Authorized 
representatives as noted above are bound to maintain the strict confidentiality of medical 
and research information that may be linked to identified individuals. The investigational 
sites will normally be notified in advance of auditing visits. 
11. 
QUALITY CONTROL AND QUALITY ASSURANCE 
The principal investigator is required to keep accurate records to ensure that the conduct 
of the study is fully documented. The principal investigator is required to ensure that all 
eCRFs are completed for every participant entered in the trial. 
The DAIT/NIAID is responsible for regular inspection of the conduct of the study, for 
verifying adherence to the protocol, and for confirming the completeness, consistency, 
and accuracy of all documented data. 
The eCRFs will be completed online via a web-based electronic data capture (EDC) 
system that has been validated and is compliant with Part 11 Title 21 of the Code of 
Federal Regulations. Some data requirements will be addressed outside the EDC using 
SAS® software. Data queries will be issued and resolved within the EDC system or 
SAS®. 
Study staff at the site will enter information into the electronic CRFs, and the data will be 
stored remotely at a central database. Data quality will be ensured through the EDC 
system’s continuous monitoring of data and real-time detection and correction of errors. 
All elements of data entry (i.e., time, date, verbatim text, and the name of the person 
performing the data entry) will be recorded in an electronic audit trail to allow all 
changes in the database to be monitored and maintained in accordance with federal 
regulations.  
Study staff will enter data from a study visit on the relevant eCRFs within 3 days 
following the visit or the time when data become available. 
12. 
ETHICAL CONSIDERATIONS AND COMPLIANCE WITH GOOD CLINICAL 
PRACTICE 
12.1 
STATEMENT OF COMPLIANCE 
This trial will be conducted in compliance with the protocol, current Good Clinical 
Practice (GCP) guidelines—adopting the principles of the Declaration of Helsinki—and 
all applicable regulatory requirements. 
Prior to study initiation, the protocol and the informed consent documents will be 
reviewed and approved by the sponsor and the Ethics Committee. Any amendments to 
the protocol or consent materials must also be approved by the sponsor and Ethics 
Committee before they are implemented. 

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12.2 
INFORMED CONSENT 
The informed consent form is a means of providing information about the trial to a 
prospective participant and allows for an informed decision about participation in the 
study. For participants (or their legally acceptable representative) attending a visit in 
person, they must read, sign, and date a consent form before participating in the study, 
and/or undergoing any study-specific procedures. If a participant does not speak and read 
English, the consent materials must be translated into the appropriate language. 
The informed consent form must be updated or revised whenever important new safety 
information is available, whenever the protocol is amended, and/or whenever any new 
information becomes available that may affect participation in the trial. 
A copy of the informed consent will be given to a prospective participant for review. A 
clinical team member will review the consent and answer questions. The participant will be 
informed that participation is voluntary and that he/she may withdraw from the study at 
any time, for any reason. 
Families of former LEAP participants will be sent a detailed Patient Information Sheet 
(including age-appropriate versions for children) and Informed Consent Form shortly 
before their study visit. At the beginning of the visit a clinical team member will re-
discuss the study, explaining it to the family's children in an age-appropriate way and 
taking assent from children younger than 16 years of age. Persons with parental 
responsibility will then sign an individual consent form for each participating family 
member before any procedures take place. Appropriately aged siblings will sign their 
own consent. For families who agree to data collection via questionnaire only, a 
completed questionnaire will be considered as implicit consent. 
For families who agree to data collection via questionnaire only, a returned completed 
questionnaire will be considered as implicit consent. For participants completing a 
telephone visit only, participants will be asked to give verbal consent before any visit 
data is collected. In advance of the call families/parents of former LEAP participants will 
be sent the Patient Information Sheet and Informed Consent Form. The information will 
be sent prior to the telephone visit to give participants time to consider participation and 
to ask questions. At the beginning of the telephone visit a clinical team member will 
review the study with a person with parental responsibility and provide an opportunity to 
ask questions before taking and noting verbal consent. 
12.3 
PRIVACY AND CONFIDENTIALITY 
A participant’s privacy and confidentiality will be respected throughout the study. Each 
participant will be assigned a sequential identification number. This number, rather than 
the participant’s name, will be used to collect, store, and report participant information. 
13. 
PUBLICATION POLICY 
The ITN policy on publication of study results will apply to this study. Authorized 
participants may find details regarding the policy statement on the ITN internet website at 
http://www.immunetolerance.org.  

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14. 
REFERENCES 
1. Sicherer SH, Munoz-Furlong A, Godbold JH, Sampson HA. US prevalence of self-reported peanut, 
tree nut, and sesame allergy: 11-year follow-up. J Allergy Clin Immunol 2010;125:1322-6. 
2. Boyce JA, Assa'ad A, Burks AW, et al. Guidelines for the diagnosis and management of food allergy 
in the United States: report of the NIAID-sponsored expert panel. J Allergy Clin Immunol 
2010;126:S1-58. 
3. Du Toit G, Roberts G, Sayre PH, et al. Randomized trial of peanut consumption in infants at risk for 
peanut allergy. N Engl J Med 2015;372:803-13. 
4. Perkin MR, Logan K, Marrs T, et al. Enquiring About Tolerance (EAT) study: Feasibility of an early 
allergenic food introduction regimen. J Allergy Clin Immunol 2016;137:1477-86 e8. 
5. Du Toit G, Sayre PH, Roberts G, et al. Effect of Avoidance on Peanut Allergy after Early Peanut 
Consumption. N Engl J Med 2016;374:1435-43. 
6. Togias A, Cooper SF, Acebal ML, et al. Addendum guidelines for the prevention of peanut allergy in 
the United States: Report of the National Institute of Allergy and Infectious Diseases-sponsored expert 
panel. J Allergy Clin Immunol 2017;139:29-44. 
7. Feeney M, Du Toit G, Roberts G, et al. Impact of peanut consumption in the LEAP Study: Feasibility, 
growth, and nutrition. J Allergy Clin Immunol 2016. 
8. Liem JJ, Huq S, Kozyrskyj AL, Becker AB. Should Younger Siblings of Peanut-Allergic Children Be 
Assessed by an Allergist before Being Fed Peanut? Allergy Asthma Clin Immunol 2008;4:144-9. 
9. Sicherer SH, Furlong TJ, Maes HH, Desnick RJ, Sampson HA, Gelb BD. Genetics of peanut allergy: a 
twin study. J Allergy Clin Immunol 2000;106:53-6. 
10. Hong X, Hao K, Ladd-Acosta C, et al. Genome-wide association study identifies peanut allergy-
specific loci and evidence of epigenetic mediation in US children. Nat Commun 2015;6:6304. 
11. Baye TM, Martin LJ, Khurana Hershey GK. Application of genetic/genomic approaches to allergic 
disorders. J Allergy Clin Immunol 2010;126:425-36; quiz 37-8. 
12. Du Toit G, Roberts G, Sayre PH, et al. Identifying infants at high risk of peanut allergy: the Learning 
Early About Peanut Allergy (LEAP) screening study. The Journal of allergy and clinical immunology 
2013;131:135-43 e1-12. 
13. Turcanu V, Maleki SJ, Lack G. Characterization of lymphocyte responses to peanuts in normal 
children, peanut-allergic children, and allergic children who acquired tolerance to peanuts. J Clin 
Invest 2003;111:1065-72. 
14. Flinterman AE, Pasmans SG, den Hartog Jager CF, et al. T cell responses to major peanut allergens in 
children with and without peanut allergy. Clin Exp Allergy 2010;40:590-7. 
15. DeLong JH, Simpson KH, Wambre E, James EA, Robinson D, Kwok WW. Ara h 1-reactive T cells in 
individuals with peanut allergy. J Allergy Clin Immunol 2011;127:1211-8.e3. 
16. Wambre E, DeLong JH, James EA, LaFond RE, Robinson D, Kwok WW. Differentiation stage 
determines pathologic and protective allergen-specific CD4+ T-cell outcomes during specific 
immunotherapy. J Allergy Clin Immunol 2012;129:544-51, 51.e1-7. 
17. Flinterman AE, Knol EF, Lencer DA, et al. Peanut epitopes for IgE and IgG4 in peanut-sensitized 
children in relation to severity of peanut allergy. J Allergy Clin Immunol 2008;121:737-43.e10. 
18. Shamji MH, Wilcock LK, Wachholz PA, et al. The IgE-facilitated allergen binding (FAB) assay: 
validation of a novel flow-cytometric based method for the detection of inhibitory antibody responses. 
Journal of immunological methods 2006;317:71-9. 
19. Vickery BP, Scurlock AM, Kulis M, et al. Sustained unresponsiveness to peanut in subjects who have 
completed peanut oral immunotherapy. J Allergy Clin Immunol 2014;133:468-75. 
20. Santos AF, James LK, Bahnson HT, et al. IgG4 inhibits peanut-induced basophil and mast cell 
activation in peanut-tolerant children sensitized to peanut major allergens. J Allergy Clin Immunol 
2015;135:1249-56. 
21. Santos AF, Douiri A, Becares N, et al. Basophil activation test discriminates between allergy and 
tolerance in peanut-sensitized children. J Allergy Clin Immunol 2014;134:645-52. 
 

Immune Tolerance Network 
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LEAP Trio, IRAS ID 195451 
APPENDIX 1. SCHEDULE OF EVENTS 
V 1441 
LEAP Participant 
Parents 
Siblings 
General Assessments 
 Informed consent 
x 
x 
x 
Medical history 
x 
x 
x 
Physical examination2 
x 
 
x 
Pregnancy test3 
x 
 
x 
Concomitant medications 
x 
x 
x 
Eczema - SCORAD 
x 
 
x 
Skin - TEWL 
x 
 
x 
Rhinitis  & OAS evaluation 
x 
x 
x 
Asthma – Spirometry 
x 
 
x 
Food reaction history 
x 
x 
x 
Adverse events 
x 
 
x 
Dietary Assessments/Exposure 
Three-day food diary 
x 
 
 
Peanut consumption 
monitoring  
x 
x 
x 
Bed dust collection 
x 
 
x 
Dietary education 
x 
 
x 
Allergy Assessments 
SPT for peanut 
x 
x 
x 
SPT for ingested allergens 
x 
x 
x 
SPT for inhalant allergens 
x 
x 
x 
IgE total4 
x 
x 
x 
IgE and IgE CRD for peanut 
x 
x 
x 
IgG for peanut 
x 
x 
x 
IgG4 and IgG4 CRD for 
peanut 
x 
x 
x 
                                                          
1 Home visits or telephone visits may be completed for participants who are unable to attend the clinical 
trial unit. Sections 6.2.2 and 6.2.3 provide additional details regarding assessments to be completed at 
these visits. 
2 Temperature, pulse, respiratory rate, weight and height will be completed for LEAP participants and 
siblings <18 years of age. Skin fold thickness and waist circumference measurements will only be taken 
for LEAP participants. 
3 A urine based pregnancy test is required for all LEAP participants and siblings who are post-menarchal 
and who will undergo an incremental peanut oral food challenge. 
4 For participants who are unable to attend the clinical trial unit, a home visit or home blood collection is 
not possible, IgE results will be accepted for those undertaken in standardized labs using Thermo Fisher 
Scientific ImmunoCap System. 

Immune Tolerance Network 
CONFIDENTIAL 
Page 54 
Protocol ITN070AD 
v 7.0  
10 December 2021 
LEAP Trio, IRAS ID 195451 
V 1441 
LEAP Participant 
Parents 
Siblings 
IgE for ingested allergens 
x 
x 
x 
IgE for inhalant allergens 
x 
x 
x 
Oral peanut food challenges 
x 
 
x5 
 
 
 
 
Mechanistic Assessments 
PBMC cellular assays 
x 
x 
x 
Plasma archive 
x 
x 
x 
Serum archive 
x 
x 
x 
Whole blood DNA/HLA 
genotype 
x 
x 
x 
Whole blood RNA 
x 
 
x 
 
 
 
 
 
 
 
 
 
 
 
                                                          
5 Oral peanut food challenges will only be performed in siblings <18 years of age. 

Immune Tolerance Network 
CONFIDENTIAL 
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Protocol ITN070AD 
v 7.0  
10 December 2021 
LEAP Trio, IRAS ID 195451 
 
 
APPENDIX 2. PREVIOUSLY USED ALGORITHM FOR DETERMINING PEANUT 
ALLERGY STATUS 
Prior to development of peanut allergy prediction, the algorithm below was used to determine 
peanut allergy status when allergic status could not be determined by history or oral food 
challenge. Features of this algorithm, which is based on the one used in the LEAP Study,3 are 
shown in Figure 3. 
For purposes of this algorithm, trace exposure to peanut is considered consumption of 0.25 g 
peanut protein in a single exposure. Low range IgE is defined as <0.35 kU/L; mid-range is 
defined as > 0.35 and <15 and high range is defined as ≥15 kU/L. 
1. Individuals with more than trace exposure and no symptoms will be  
• regarded as tolerant if they have  
SPT <3 mm and IgE in the low range or no results for IgE; or  
if they have no data for SPT and have IgE in low range. 
2. Individuals with less than or equal to trace exposure will be  
• regarded as tolerant if they have 
SPT <3 mm, and IgE in the low range or no data for IgE; or  
no data for SPT and IgE in the low range.  
• regarded as allergic if they have 
SPT 3-7 mm inclusive and IgE in the high range; 
SPT ≥8 mm and IgE in the mid-range, high range or no data; or 
no data for SPT and IgE in the high range. 
3. Individuals with symptoms related to exposure will be  
• regarded as allergic if they have 
SPT 3-7 mm inclusive and IgE in the high range; 
SPT ≥8 mm and IgE in the mid-range, high range or no data; or 
no data for SPT and IgE in the high range. 
4. Individuals who report eating peanut regularly, will not be regarded as allergic independent of 
other results. 
Additional combinations of outcomes are in Figure 5. 

Immune Tolerance Network 
CONFIDENTIAL 
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Protocol ITN070AD 
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LEAP Trio, IRAS ID 195451 
Peanut exposure
and symptoms
SPT
(mm)
IgE
Mid or
ND
<3
3–7
>8
No 
data
N
IgE
Mid or
ND
N
IgE
Mid or
ND
Al
IgE
Mid or
ND
N
SPT
(mm)
IgE
Mid
<3
3–7
>8
No 
data
N
IgE
Mid or
ND
N
IgE
Mid or
ND
Al
IgE
Mid or
ND
N
IgE
Mid
<3
3–7
>8
No 
data
N
IgE
Mid or
ND
N
IgE
Mid or
ND
N
IgE
Mid or
ND
N
Nonevaluable
N
N
N
Al
N
Al
N
Al
Tol
N
N
N
N
N
Tol
N
Tol
N
N
Al
N
Al
Tol
Al
SPT
(mm)
IgE Levels in kU/L
Low:  < 0.35
Mid:  > 0.35 and < 15
Hi:    >15
ND:   No data
Allergic status
Tol:  tolerant
Al:   allergic
N:    nonevaluable
Trace is defined as 0.25 g peanut 
protein in a single exposure
More than trace exposure
No symptoms
Less than or equal to trace exposure
Symptoms related to 
exposure
UNKNOWN TENDANCY
TOLERANT TENDENCY
ALLERGIC TENDENCY
No data
 
Figure 5. Prior algorithm for determination of peanut allergy in individuals who do not 
undergo an oral food challenge 
This algorithm is to be replaced by the peanut allergy prediction model. This algorithm begins with an 
assessment of peanut exposure and related symptoms. If no data regarding this is available, the subject is 
non-evaluable by this algorithm. Otherwise, they will be evaluated as described in the text.