A Prospective, Vehicle Controlled, Double Blind, Multicenter, Randomized, Phase II Study of B244 Delivered as a Topical Spray to Determine Safety and Efficacy in Subjects With Mild to Moderate Atopic Dermatitis
Phase: PHASE2
Conditions: Atopic Dermatitis Eczema
Sponsor: AOBiome LLC

CONFIDENTIAL 
 
Amendment 3 
 
1 
 
 
 
CLINICAL TRIAL PROTOCOL 
Phase II 
 
COMPOUND: 
B244 
 
 
 
 
A Prospective, Vehicle Controlled, Double Blind, Multicenter, Randomized, Phase 
II Study of B244 Delivered as a Topical Spray to Determine Safety in Subjects with 
Mild to Moderate Atopic Dermatitis. 
 
 
STUDY NUMBER: ADB244-001 
 
 
 
VERSION DATE: October 5, 2018 
 
 
 
 
Sponsor: 
AOBiome Therapeutics 
125 Cambridgepark Drive 
Cambridge, MA 02140 
 
 
 
Any and all information presented in this document shall be treated as confidential and shall remain the 
exclusive property of AOBiome Therapeutics (or any of its affiliated companies). The use of such 
confidential information must be restricted to the recipient for the agreed purpose and must not be 
disclosed, published or otherwise communicated to any unauthorized persons, for any reasons, in any form 
whatsoever without the prior written consent of AOBiome Therapeutics (or the concerned affiliated 
company)
NCT03235024

CONFIDENTIAL 
 
Amendment 3 
 
2 
 
Revision Chronology 
 
Original 
June 14, 2017 
Amendment 1 
September 1, 2017 
Amendment 2 
January 10, 2018 
Amendment 3 
October 5, 2018 
 
 
 

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Amendment 3 
 
4 
INVESTIGATOR AGREEMENT 
 
I have read this protocol and agree: 
• 
To conduct the study as outlined herein, in accordance with Good Clinical 
Practices (GCPs), the Declaration of Helsinki and complying with the obligations 
and requirements of Clinical Investigators and all other requirements listed in 21 
CFR part 312 and according to the study procedures provided by AOBiome LLC 
and local regulations. 
• 
Not to implement any changes to the protocol without prior agreement from the 
Sponsor and prior review and written approval from the IRB or IEC, except as 
would be necessary to eliminate an immediate hazard to study participant (s), or 
for administrative aspects of the study. 
• 
To ensure that all persons assisting me with the study are adequately informed 
about the investigational product(s) and of their study-related duties as described 
in the protocol. 
• 
To completely inform all participants in this study concerning the pertinent details 
and purpose of the study prior to their agreement to participate in the study in 
accordance with GCP and regulatory authority requirements. 
• 
To be responsible for maintaining each participant’s consent form in a secure 
study file and providing each participant with a signed copy of the consent form. 
• 
That I am thoroughly familiar with the appropriate use of the investigational 
product(s), as described in the protocol, and any additional information provided 
to me by, or on behalf of AOBiome Therapeutics 
 
 
 
Investigator Printed Name:  
 
 
 
 
 
 
 
 
 
 
 
 
 
Signature: 
 
 
 
 
 
Date:   
 
 
 
 
 
 
 

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Amendment 3 
 
Page 5  
Statement of Compliance 
 
 
The study will be carried out in accordance with Good Clinical Practice (GCP) as required by the 
following: 
● United States (US) Code of Federal Regulations (CFR) applicable to clinical studies (45 
CFR Part 46; 21 CFR Part 50, 21 CFR Part 56, and 21 CFR Part 312) 
● ICH E6; 62 Federal Register 25691 (1997) 
 
All key personnel (all individuals responsible for the design and conduct of this study) have 
completed Good Clinical Practice Training. 
 
 

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Amendment 3 
 
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PROTOCOL AMENDMENT 3 SUMMARY OF CHANGES 
 
Section 
Description of Changes 
Title Page 
Updated with Sponsor’s new company logo and 
business address 
Sponsor Approval Page 
Updated section with up to date information 
Clinical Trial Summary 
Section 6.2 
Per Administrative Letter dated 27FEB2018, 
Inclusion criteria for the study has been updated as 
follows: increase the upper limit of subject’s BSA 
affected by Atopic Dermatitis to 30%. 
Section 11.1 
Schedule of Events Table 
Per Administrative Letter dated 12MAR2018: 
inserted clarification that that medical records 
from patient’s dermatologist or primary care 
physician to confirm the diagnosis are optional. 
Verbal confirmation of the diagnosis present for ³ 
12 months is sufficient in order to fulfill this 
criterion.  
Sections 7.2; 8.15.1; 11.6.2;  
Schedule of Events Table 
 
Per Administrative Letter dated 26APR2018, 
inserted clarification language stating that the use 
of moisturizer in the study is not mandatory. 
Sections 9; 11.6; 12.11 
Schedule of Events Table 
Per Administrative Letter dated 17JUL2018, 
subjects will be provided with two (2) actigraphy 
watches at Baseline period and will be asked to 
wear one watch on each wrist at all times through 
Day 28. Subjects will be asked to return both 
watches at Day 28 visit.  
 
Section 8.7 
Modified language around IP kits 
Section 12.1 
Updated shipping schedule and corrected Lab 
name 
Section 12.11 
Updated Actigraphy watch schedule and 
procedures 
Section 12.14 
Revised language around sample shipment to state 
that samples will be shipped according to the Lab 
Manual.  
Section 16.7 
Updated safety reporting email 
 
Administrative changes: Minor changes involving grammar, wordsmithing, punctuation, and 
other editorial changes have been made throughout the document. All are clearly identified in the 
track-changes version of the amendment. 
 
Additional formatting and stylistic adjustments have been made to facilitate the reading 
process. 
 
 
 

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CLINICAL TRIAL SUMMARY 
 
COMPOUND: B244  
 
 
 
STUDY NUMBER: ADB244-001 
TITLE: 
A Prospective, Vehicle Controlled, 
Double Blind, Multicenter, Randomized, 
Phase II Study of B244 Delivered as a 
Topical Spray to Determine Safety in 
Subjects with Mild to Moderate Atopic 
Dermatitis. 
INVESTIGATIONAL PRODUCT 
B244 
 
Topical application 
STUDY ARMS 
1. B244  
2. Vehicle 
PURPOSE: 
The aim of the study is to assess the safety 
of B244 in the treatment of atopic 
dermatitis. 
 
STUDY OBJECTIVES: 
Primary:  
• To evaluate the safety and 
tolerability of B244 in participants 
with mild to moderate Atopic 
Dermatitis. 
Secondary:  
• To assess the efficacy of B244 
versus vehicle by the change in 
Atopic Dermatitis Area Severity 
Index (EASI) Score from Baseline 
to post-baseline visits 
 
 Exploratory: 

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Amendment 3 
 
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• To assess the efficacy of B244 
versus vehicle by the change in 
Visual Analog Scale (VAS) Score 
from Baseline to post-baseline 
visits 
• To assess the efficacy of B244 
versus vehicle by the change in 
Skindex 16 Score from Baseline to 
post-baseline visits 
• To assess the efficacy of B244 
versus vehicle by the change in 
Investigator Global Assessment 
Score (IGA) from Baseline to 
post-baseline visits 
• To assess the efficacy of B244 
versus vehicle by the change in 
Actigraphy Movement Count per 
Hour and sleep quality during 
nighttime from Baseline to post-
baseline visits 
• To evaluate if B244 administration 
on the skin twice daily for 28 days 
will affect the levels of immune 
biomarkers. 
• To explore microbial content at 
baseline and Day 28. 
 
STUDY DESIGN: 
This is a Prospective, Vehicle Controlled, 
Double Blind, Multicenter, Randomized 
Phase II trial, comparing the effect of 
twice daily B244 application for 28 days 
vs vehicle application on treatment of 
mild to moderate AD  
At Screening and Baseline, all subjects 
must have atopic dermatitis, as defined by 
the Hanifin and Rajka criteria, which 
involves a minimum of 10% and a 
maximum of 30% body surface area, 
EASI score of 10 to 21 and pruritus visual 
analogue scale scores of ≥ 5 points on the 

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VAS scale (at least moderate), IGA of 2-
3. 
The total duration of the study will be 
approximately 9 weeks. Participants will 
report for a Screening visit and if all 
inclusion criteria are met, subjects will go 
through a two week washout phase before 
reporting for a Baseline visit.  
Subjects will come in for visits at Day 14 
(Week 2), Day 28 (Week 4). A final visit 
will be conducted at Day 42 (Week 6).   
Efficacy will be assessed using Atopic 
Dermatitis Area and Severity Index 
(EASI), Visual Analog Scale (VAS) and 
Investigator Global Assessment (IGA) 
score .  
Blood and urine samples will be collected 
for standard safety laboratory tests and 
effect of the drug on inflammatory 
biomarkers. Participant's safety will be 
monitored throughout the study. 
We plan to enroll approximately 130 total 
patients. Assuming 20% drop out rate, we 
will plan to complete 104 patients.  
Randomization will be 1:1 so that equal 
number of patients will be treated in each 
Arm of the study. 
All B244 randomized subjects will be 
treated at the dose of 4x109 cfu/ml (mid-
dose of the Phase 1b/2a safety trial) 
STUDY POPULATION: 
 
Male and female subjects 18 years of age 
or older with clinically diagnosed mild to 
moderate Atopic Dermatitis.   
Main Inclusion Criteria: 
 
• Male and female subjects ≥18 
years of age 

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Amendment 3 
 
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• In good general health as 
determined by a thorough medical 
history and physical examination, 
and vital signs 
• Clinical diagnosis of mild to 
moderate atopic dermatitis 
according to the criteria of Hanifin 
and Rajka  
• Mild to moderate Atopic 
Dermatitis area and severity index 
[EASI] 10-21 
• A score of at least ≥ 5 points  
(moderate pruritus) on the VAS 
for pruritus 
• A minimum of 10% and not more 
than 30% of the subjects' BSA 
affected by atopic dermatitis 
(affected is defined by physical 
examination findings: erythema, 
edema, scaling, lichenification, 
excoriation, with the excoriation 
serving as the physical 
examination correlate of pruritus) 
• An IGA score of 2-3 
• Patient has a history of AD for ³ 
12 months 
• Ability to read and understand 
English and to provide written 
informed consent and 
authorization for protected health 
information disclosure 
 
Main Exclusion Criteria: 
 
 
• Pregnant and lactating women by 
urine pregnancy testing 
• Subjects with any significant 
clinical abnormalities which may 
interfere with study participation 
• Any skin condition which may 
interfere with evaluation of AD 
• Atopic dermatitis on the head or 
scalp 

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Amendment 3 
 
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• Subjects with Atopic dermatitis on 
the face 
• Unstable or actively infected 
atopic dermatitis 
• Patients suffering from pruritus 
from conditions other than AD 
• Patients with chronic pruritus due 
to systemic disease  
• Patients with conditions requiring 
inhaled steroids 
• Have concurrent skin disease of 
such severity in the study area that 
it could interfere with the study 
evaluation 
• Have active skin infections on the 
treatment area 
• Have received or planning to 
receive topical corticosteroids, 
topical coal tar, topical sulfur, 
topical PDE-4 inhibitors, topical 
antihistamines, topical antiseptics 
or antibiotics, topical antifungals, 
bleach baths, UVA or UVB 
phototherapy, oral/IV/inhaled 
steroids,  
antibiotics/antiviral/antifungal 
agents, oral probiotics, 
glucocorticoids treatment, 
calcineurin inhibitors, 
immunomodulating biologic 
agents, systemic glucocorticoids, 
systemic immunosuppressive or 
immunomodulatory agents within 
2 weeks of Baseline visit.   
• Current or recent history (£ 3 
months of systemic use of 
Otrexup™, Rasuvo®, 
Rheumatrex® and Trexall™ or its 
generic versions such as 
Methotrexate  
• History of being seropositive for 
human immunodeficiency virus 
(HIV) at screening by laboratory 
testing at Screening 

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• History of being positive for 
Hepatitis B virus surface antigen 
(HBsAg) or positive Hepatitis C 
virus antibody (HCV Ab) at 
screening by laboratory testing at 
Screening 
• History of renal disease 
• Use of any investigational drugs 
within the previous 30 days prior 
to dosing or within a period of less 
than five times the drug's half-life, 
whichever is longer 
• Use of any biologic within a 
period of 5 times its half-life 
• Use of vinegar or bleach baths 
within 2 weeks of starting the 
study 
 
DOSE REGIMEN: 
Subjects will apply a total of 8 pumps of 
IP per application to all affected 
areas twice-a-day (i.e. 8 pumps in the 
morning and 8 pumps again at night) for 
28 Days. 
 
ASSESSMENT SCHEDULE: 
All subjects will attend a screening visit 
not more than 21 days prior to Day 1. 
Subjects will be required to return to the 
clinic at Baseline, on Day 14 (Week 2), 
Day 28 (Week 4). All subjects will be 
asked to attend a follow-up visit 2 weeks 
(14 (±3) days) after the last dose of study 
medication. 
STATISTICAL CONSIDERATIONS: 
• All efficacy analyses will be 
performed on the ITT population 
and PP population  
• The difference in treatment groups 
in change-from-baseline values at 
post-baseline visits will be 
analyzed using a mixed Analysis 
of Variance with one between-
subject variable (active group vs. 
vehicle group) and one within-

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subject variable 
• 
One-sided two-sample 
equal-variance t-test will be 
performed as a supportive test to 
analyze the differences in 
treatment group means in change 
from baseline at each post-baseline 
visit  
 
DURATION OF STUDY: 
Subject’s participation in the study will 
take up to 9 weeks. The estimated total 
study duration is 12 months. All subjects 
will be followed from enrollment and until 
the Final (End of Study Visit), which 
occurs 42 days following the baseline 
visit. 

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1 
STUDY SCHEMA 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Male and Female subjects with clinical diagnosis of Atopic 
Dermatitis (AD) 
Vehicle 
Treatment and Study Assessments at: 
Baseline, Week  2, 4 
B244 
End of Study visit at Week 6 
 
Screening visit 
-21 to -14 
Washout phase 
-14 to -1 
Baseline visit 
-1 to 0 
Randomize 

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Table of Contents 
1 
STUDY SCHEMA ................................................................................................................ 14 
Table of Contents ..................................................................................................................... 15 
2 
INTRODUCTION ................................................................................................................ 20 
2.1 
Background .......................................................................................................................... 20 
3 
STUDY OBJECTIVES ........................................................................................................... 21 
3.1 
Primary Objectives ............................................................................................................... 21 
3.2 
Secondary Objectives ........................................................................................................... 21 
3.3 
Exploratory Objectives ......................................................................................................... 21 
4 
ENDPOINTS ...................................................................................................................... 22 
4.1 
Safety & tolerability: ............................................................................................................ 22 
4.2 
Efficacy:................................................................................................................................ 22 
4.3 
Exploratory: ......................................................................................................................... 22 
5 
STUDY DESIGN .................................................................................................................. 22 
6 
SELECTION OF STUDY PARTICIPANTS ............................................................................... 23 
6.1 
Number of Participants Planned .......................................................................................... 23 
6.2 
Inclusion Criteria .................................................................................................................. 23 
6.3 
Exclusion Criteria ................................................................................................................. 23 
7 
PARTICIPANT ENROLMENT............................................................................................... 24 
7.1 
Consenting Participants ....................................................................................................... 24 
7.2 
Screening for Eligibility ........................................................................................................ 24 
7.3 
Study Withdrawal and Withdrawal From Investigational Product and Stopping Criteria .... 25 
7.4 
Screen Failures ..................................................................................................................... 25 
7.5 
Early Termination ................................................................................................................ 25 
8 
STUDY TREATMENT .......................................................................................................... 25 
8.1 
Investigational Product ........................................................................................................ 25 
8.2 
Dose Changes ....................................................................................................................... 26 
8.3 
Storage conditions ............................................................................................................... 27 
8.4 
Description of Blinding Method ........................................................................................... 27 
8.5 
Treatment Assignments: ...................................................................................................... 27 
8.6 
Treatment Compliance......................................................................................................... 27 
8.7 
Treatment Application ......................................................................................................... 27 
8.8 
Treatment of Investigational Product Overdose .................................................................. 28 
8.9 
Product Accountability......................................................................................................... 28 
8.10 
Unblinding Procedures ......................................................................................................... 28 
8.11 
Retrieval and Destruction of Investigational Product .......................................................... 28 
8.12 
Permitted Medications ........................................................................................................ 29 
8.13 
Prohibited Medications........................................................................................................ 29 
8.14 
Rescue Medications ............................................................................................................. 29 
8.15 
Lifestyle Restrictions ............................................................................................................ 29 
8.15.1 
Use of cleanser and moisturizer during the trial ........................................................... 29 

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8.16 
Handling of Investigational Product ..................................................................................... 30 
8.17 
Treatment compliance ......................................................................................................... 30 
9 
Actigraphy Watch ............................................................................................................. 30 
10 
CONTRACEPTION REQUIREMENTS ............................................................................... 30 
11 
STUDY PROCEDURES .................................................................................................... 31 
11.1 
Pre-screening procedures .................................................................................................... 31 
11.2 
Informed Consent Procedures ............................................................................................. 31 
11.3 
Study Assessments............................................................................................................... 31 
11.4 
Inclusion procedures ............................................................................................................ 32 
11.5 
Timing of patient’s visits to the clinic ................................................................................... 32 
11.6 
Description by type of visit .................................................................................................. 32 
11.6.1 
Screening Visit (-21 to -14) ........................................................................................... 32 
11.6.2 
Washout Phase (-14 to -1) ............................................................................................ 33 
11.6.3 
Study Day 1-Baseline visit (-1 to 0)................................................................................ 33 
11.6.4 
Week 2, 4 (Day 14, 28) study visit ................................................................................. 33 
11.6.5 
Week 6 final visit (Day 42)-End of Study Visit ................................................................ 34 
11.6.6 
Unscheduled/Unanticipated Study visit ........................................................................ 34 
11.6.7 
Early Termination Visit ................................................................................................. 35 
12 
Methods of Assessments .............................................................................................. 35 
12.1 
Skin Swab Samples............................................................................................................... 35 
12.2 
Blood Pressure Measurement .............................................................................................. 36 
12.3 
Physical Examinations .......................................................................................................... 36 
12.4 
Laboratory Assessments ...................................................................................................... 36 
12.5 
Oral Hygiene Questions ....................................................................................................... 37 
12.6 
EASI Assessment .................................................................................................................. 37 
12.7 
Subject reported outcomes/Skindex16 ................................................................................ 37 
12.8 
Hanifin and Rajka Diagnostic Criteria for Atopic Dermatitis (AD) ......................................... 37 
12.9 
Visual Analog Scale (VAS)..................................................................................................... 38 
12.10 
Investigator Global Assessment Score (IGA) .................................................................... 39 
12.11 
Actigraphy Watch ............................................................................................................ 39 
12.12 
Biomarkers ...................................................................................................................... 40 
12.13 
Microbial Content ............................................................................................................ 40 
12.14 
Sample Shipment ............................................................................................................. 40 
13 
Safety Assessments ...................................................................................................... 41 
13.1 
Compliance .......................................................................................................................... 41 
13.2 
Pregnancy Reporting ............................................................................................................ 41 
13.3 
Study Completion ................................................................................................................ 41 
13.4 
Subject Withdrawal Criteria ................................................................................................. 41 
14 
EFFICACY ASSESSMENTS ............................................................................................... 42 
15 
STATISTICAL CONSIDERATIONS .................................................................................... 42 
15.1 
Sample Size .......................................................................................................................... 42 
15.2 
Populations for Analysis ...................................................................................................... 42 
15.3 
Data Analysis ....................................................................................................................... 42 

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15.3.1 
Disposition ................................................................................................................... 43 
15.3.2 
Demographic and Baseline ........................................................................................... 43 
15.4 
Safety Analyses .................................................................................................................... 43 
15.4.1 
Definitions.................................................................................................................... 43 
15.4.2 
Adverse Events ............................................................................................................. 43 
15.4.3 
Deaths and Serious Adverse Events .............................................................................. 43 
15.4.4 
Adverse Events leading to treatment discontinuation ................................................... 43 
15.4.5 
Efficacy Analyses .......................................................................................................... 44 
15.4.6 
Handling of dropouts or missing data ........................................................................... 44 
15.5 
Clinical Trial Protocol deviations .......................................................................................... 44 
16 
ADVERSE EVENTS (AE) AND SERIOUS ADVERSE EVENTS (sae) ...................................... 44 
16.1 
Definition of an AE ............................................................................................................... 44 
16.2 
Definition of a SAE ............................................................................................................... 45 
16.3 
Time Period, Frequency, and Method of Detecting AEs and SAEs ........................................ 46 
16.4 
Recording of AEs and SAEs ................................................................................................... 46 
16.5 
Evaluating AEs and SAEs ...................................................................................................... 46 
16.5.1 
Severity Rating ............................................................................................................. 46 
16.5.2 
Relationship to Investigational product (IP) .................................................................. 46 
16.6 
Pregnancy ............................................................................................................................ 47 
16.7 
Prompt Reporting of SAEs to the Sponsor ............................................................................ 47 
17 
ETHICAL AND REGULATORY STANDARDS ..................................................................... 48 
17.1 
Ethical Conduct of Study ...................................................................................................... 48 
17.2 
Laws and Regulations .......................................................................................................... 49 
17.3 
Informed Consent ................................................................................................................ 49 
17.4 
Institutional Review Board/Independent Ethics Committee (IRB/EC) .................................. 49 
17.5 
Clinical Monitoring/Record Keeping .................................................................................... 50 
18 
ADMINISTRATIVE RULES............................................................................................... 50 
18.1 
Curriculum Vitae .................................................................................................................. 50 
18.2 
Archiving of Study Documentation ...................................................................................... 50 
18.3 
Internal Safety Review Committee ...................................................................................... 50 
19 
STUDY MONITORING .................................................................................................... 51 
19.1 
Responsibilities of the Investigator(s) .................................................................................. 51 
19.2 
Responsibilities of the Sponsor ............................................................................................ 51 
19.3 
Source document requirements........................................................................................... 51 
19.4 
Use and completion of Case Report Forms (CRFs) and additional requests ......................... 52 
20 
PUBLICATIONS .............................................................................................................. 52 
21 
PROTOCOL ADHERENCE ............................................................................................... 53 
22 
CLINICAL TRIAL PROTOCOL AMENDMENTS .................................................................. 53 
23 
CONFIDENTIALITY ......................................................................................................... 53 
24 
PROPERTY RIGHTS ........................................................................................................ 53 
25 
DATA PROTECTION ....................................................................................................... 54 

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26 
SPONSOR AUDITS AND INSPECTIONS BY REGULATORY AGENCIES .............................. 54 
27 
PREMATURE DISCONTINUATION OF THE STUDY OR PREMATURE CLOSE-OUT OF A SITE
 
54 
27.1 
Decided by the Sponsor in the following cases: ................................................................... 54 
27.2 
Decided by the Investigator ................................................................................................. 55 
28 
CLINICAL TRIAL PROTOCOL AMENDMENTS .................................................................. 55 
29 
REFERENCES ................................................................................................................. 55 
30 
APPENDIX A-Schedule of Events ................................................................................... 58 
31 
Appendix B-Body Surface Area and IP application ....................................................... 62 
32 
APPENDIX C EASI QUESTIONNAIRE .............................................................................. 63 
33 
APPENDIX D. SKINDEX 16 QUESTIONNAIRE ................................................................. 64 
34 
APPENDIX E. HANIFIN AND RAJKA CRITERIA ................................................................ 65 
35 
APPENDIX F. VISAL ANALOGUE SCALE (VAS) ................................................................ 66 
36 
APPENDIX  G. IGA SCALE............................................................................................... 67 
 
 
 

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LIST OF ABBREVIATIONS AND DEFINITION OF TERMS 
 
AD 
Atopic Dermatitis 
AE 
Adverse Event 
AMO 
Ammonia Monooxygenase 
AOB 
Ammonia Oxidizing Bacteria 
BID 
Twice-Daily 
CRF 
Case Report Form 
EASI 
Atopic Dermatitis Area and Severity Index 
E/T 
Early Termination 
FDA 
Food and Drug Administration 
HAO 
NH2OH oxidoreductase 
HbsAg 
Hepatitis B Virus Surface Antigen 
HCV Ab 
Hepatitis C Virus Antibody 
HIV 
Human Immunodeficiency Virus 
ICH 
International Conference on Harmonization 
IEC 
Independent Ethics Committee 
IGA 
Investigator Global Assessment 
IP 
Investigational Product 
IRB 
Institutional Review Board 
NH2OH 
Hydroxylamine 
NH3 
Ammonia 
NO 
Nitric oxide 
NO2- 
Nitrite 
PCP 
Primary Care Physician 
SAE 
Serious Adverse Event 
SPM 
Study Procedures Manual 
VAS 
Visual Analog Scale  
 

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2 
INTRODUCTION 
2.1 
Background 
Ammonia oxidizing bacteria (AOB) are essential for the initial step in environmental nitrification processes, 
specifically the oxidation of ammonia (NH3) to nitrite (NO2-). Nitrosomonas are Gram-negative 
chemolithoautotrophic betaproteobacteria that obtain energy solely from NH3 oxidation, while fixing CO2 for 
their carbon needs.1 Oxidation of NH3 proceeds in two steps (Figure 1) leading to sequential generation of 
hydroxylamine (NH2OH) and NO2- that require two enzyme complexes: the membrane-bound ammonia 
monooxygenase (AMO) comprised of subunits AmoA, AmoB and AmoC; and the periplasmic NH2OH 
oxidoreductase (HAO). In addition to high NO2- levels, NH3 oxidation leads to nitric oxide (NO) and N2O 
production through two independent pathways downstream of NH2OH production: nitrifier denitrification and 
NH2OH oxidation.2 
 
Figure 1 
Nitrifier Denitrification Pathway 
 
B244 is a purified a strain of Nitrosomonas eutropha originally isolated from soil samples. Sequencing of the 
B244 genome revealed a distinct genetic profile from that of other published Nitrosomonas strains and AOB 
genomes. Based on in vitro co-culture studies, B244 was able to reduce survival of pathogenic bacteria. Nitrite 
generation from ammonia concurrently with medium acidification by B244 led to strong antibacterial effects 
and a marked reduction (~100-fold) in viable counts of methicillin-resistant Staphylcoccus aureus and 
Pseudomonas aeruginosa, two pathogens frequently isolated from infected skin and wound sites. In contrast, 
control cultures with B244 in the absence of ammonium or with heat-killed B244 supplemented with 
ammonium, had no antibacterial effects. The unique metabolic and antimicrobial activity of Nitrosomonas, in 
combination with their lack of virulence render these bacteria as attractive candidates for topical delivery of 
nitrite and nitric oxide on human skin with potential to improve health in both normal and abnormal skin 
conditions or wound sites. NO-releasing drugs or NO donors have also shown activity against 
Propionibacterium acnes and other pathogenic bacteria, anti-inflammatory activity, and inhibition of 
lipogenesis by insulin-stimulated immortal sebocytes.4,5  
B244 has been developed as a topical application of a natural source of AOB and NO/NOx to the human 
skin.  We hypothesize that application of AOB to eczematous skin may reduce S.aureus skin load and 
normalize the inflammatory response by reducing Th2 activation. 
  
Under IND #16487, a Phase 1b/2a clinical trial was completed  in 2016 where 36 participants with clinical 
diagnosis of facial acne vulgaris were randomized to receive twice-daily ascending doses of B244 or Vehicle 

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over 14 days. All 3 dose levels of B244 were well tolerated and no safety issues were identified.  There were no 
attributable drug related SAEs reported.  
In addition, a Phase IIb/III clinical trial in 372 patients with clinical diagnosis of facial acne is currently ongoing 
that is assessing twice-daily application of B244 or Vehicle for 3 months. To date, there have been no treatment 
related SAEs reported in this trial.  
 Eczema or Atopic dermatitis is an inflammatory skin condition that affects nearly 20% of children and 2-10% 
of adults20. Disease prevalence has steadily grown in the last 30 years, resulting in a growing field of atopic 
dermatitis reasearch21 
The disease is characterized by pruritus, scratching, and eczematous lesions. The chronic condition may be 
associated with thickening and pigmentary changes. Onset of the disease occurs mostly between 3 and 6 
months, but in 90% of children, the disease occurs by age of 5. Staphylococcus aureus is consistently found in 
eczematous skin lesions in patients with AD. Correlation between the severity of the disease and presence 
of Staphylococcus aureus has been well established and it has been shown that presence of bacteria is an 
important factor in skin aggravation22. The goal of therapy for AD is to restore the epidermal barrier function 
and reduce skin inflammation. However, systemic antibiotic use is controversial.  
 The primary purpose of this study is to evaluate the safety of B244 in treating patients with Atopic Dermatitis. 
3 
STUDY OBJECTIVES 
3.1 
Primary Objectives 
• To evaluate the safety and tolerability of B244 in participants with mild to moderate Atopic Dermatitis.  
3.2 
Secondary Objectives 
• To assess the efficacy of B244 versus vehicle by the change in Atopic Dermatitis Area Severity Index 
(EASI) Score from Baseline to post-baseline visits 
 
3.3 
Exploratory Objectives 
• To assess the efficacy of B244 versus vehicle by the change in Visual Analog Scale (VAS) Score from 
Baseline to post-baseline visits 
• To assess the efficacy of B244 versus vehicle by the change in Skindex 16 Score from Baseline to post-
baseline visits 
• 
To assess the efficacy of B244 versus vehicle by observing a change in the IGA score from Baseline to 
post-baseline visits 
 
• To assess the efficacy of B244 versus vehicle by the change in Actigraphy Movement Count per Hour 
and sleep quality during nighttime from Baseline to post-baseline visits 
• To evaluate if B244 administration on the skin twice daily for 28 days will affect the levels of immune 
biomarkers. 
• To explore microbial content at baseline and Day 28.  

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4 
ENDPOINTS 
4.1 
Safety & tolerability: 
• Safety and tolerability endpoints will consist of all adverse events reporting during the study duration. 
4.2 
Efficacy: 
• Difference in Atopic Dermatitis Area Severity Index (EASI) Score from Baseline to Day 28 
 
4.3 
Exploratory: 
• Difference in Visual Analog Scale (VAS) Score for Pruritus from Baseline to Day 28 
• Difference in the Skindex 16 Score from Baseline to Day 28 
• Difference in the IGA score from Baseline to Day 28 
• Difference in Actigraphy Movement Count per Hour and sleep quality during the night from Baseline to 
Day 28 
• Difference in biomarkers between active and vehicle groups 
• Composition of microbiome 
• Difference in skin microbiota from baseline to Day 28 between active and vehicle groups 
o Observing a difference in Staphylococcus aureus and a more diverse microbial collection on the 
patient’s skin. 
 
5 
STUDY DESIGN 
• 
This is a Prospective, Vehicle Controlled, Double Blind, Multicenter, Randomized Phase II trial, 
comparing the effect of twice daily B244 application for 28 days vs vehicle application on treatment of 
mild to moderate AD  
• 
We will enroll approximately130 patients and complete 104 patients 
• 
After screening and recruitment, participants will be randomized to active or vehicle application for 28 
days.  
• 
Subjects will apply a total of 8pumps of IP per application to all affected areas twice-a-day (i.e. 8pumps 
in the morning and 8pumps again at night) for 28 days. 
• 
Subjects will be provided with cleanser and moisturizer to use for the duration of the study 
• 
Randomization will be 1:1 so that equal number of patients will be treated in each Arm of the study. 
• 
All B244 randomized subjects will be treated at the dose of 4x109 cfu/ml 
• 
Screening will occur at Days -21 to -14 
• 
Washout will occur after Screening visit (Days -14 to -1) 
• 
Randomization will occur during the baseline period for the study  
• 
Clinical assessments of response to treatment will be made at Baseline, Study Days 14 and 28.  
• 
Subject will come back for the final visit on Study Day 42 
• 
Safety evaluations will consist of review of participant’s medical history at screening and on-going 
assessment of adverse events reported throughout the study duration. 
 

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6 
SELECTION OF STUDY PARTICIPANTS 
6.1 
Number of Participants Planned 
It is estimated that approximately 130 participants will be consented in order to provide 104 completed 
participants for randomization, treatment, and inclusion in the 
primary analysis.  
 
6.2 
Inclusion Criteria 
Deviations from inclusion criteria are not allowed because they can potentially jeopardize 
the scientific integrity of the study, regulatory acceptability or participant safety. Therefore, adherence to the 
criteria as specified in the protocol is essential. 
 
Participants eligible for enrollment in the study must meet all the following criteria: 
• Male and female subjects ≥18 years of age 
• In good general health as determined by a thorough medical history and physical examination, and vital 
signs 
• Clinical diagnosis of mild to moderate atopic dermatitis according to the criteria of Hanifin and Rajka  
• Mild to moderate Atopic Dermatitis area and severity index [EASI] 10-21 
• A score of at least  ≥ 5 points  (moderate pruritus) on the VAS for pruritus 
• A minimum of 10% and not more than 30% of the subjects' BSA affected by atopic dermatitis (affected 
is defined by physical examination findings: erythema, edema, scaling, lichenification, excoriation, with 
the excoriation serving as the physical examination correlate of pruritus)  
• An IGA score of 2-3 
• Patient has a history of AD for ³ 12 months 
• The ability to understand and sign a written informed consent form, which must be obtained prior to 
treatment 
 
6.3 
Exclusion Criteria 
Deviations from exclusion criteria are not allowed because they can potentially jeopardize the scientific 
integrity of the study, regulatory acceptability or participant safety. Therefore, adherence to the criteria as 
specified in the protocol is essential. 
 
Participants will be excluded from the study if any of the following criteria are met: 
 
• Pregnant and lactating women by urine pregnancy testing 
• Subjects with any significant clinical abnormalities which may interfere with study participation 
• Any skin condition which may interfere with evaluation of AD 
• Atopic dermatitis only on the head or scalp 
• Subjects with Atopic dermatitis on the face 
• Unstable or actively infected atopic dermatitis 
• Patients suffering from pruritus from conditions other than AD 
• Patients with chronic pruritus due to systemic disease  
• Patients with conditions requiring inhaled steroids 

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• Have concurrent skin disease of such severity in the study area that it could interfere with the study 
evaluation 
• Have active skin infections on the treatment area 
• Have received or planning to receive topical corticosteroids, topical coal tar, topical sulfur, topical PDE-
4 inhibitors, topical antihistamines, topical antiseptics or antibiotics, topical antifungals, bleach baths, 
UVA or UVB phototherapy, oral/IV/inhaled steroids,  antibiotics/antiviral/antifungal agents, oral 
probiotics, glucocorticoids treatment, calcineurin inhibitors, immunomodulating biologic agents, 
systemic glucocorticoids, systemic immunosuppressive or immunomodulatory agents within 2 weeks of 
Baseline  visit.   
• Current or recent history (£ 3 months of systemic use of Otrexup™, Rasuvo®, Rheumatrex® and 
Trexall™ or its generic versions such as Methotrexate  
• History of being seropositive for human immunodeficiency virus (HIV) at screening by laboratory 
testing at Screening 
• History of being positive for Hepatitis B virus surface antigen (HBsAg) or positive Hepatitis C virus 
antibody (HCV Ab) at screening by laboratory testing at Screening 
• History of renal disease 
• Use of any investigational drugs within the previous 30 days prior to dosing or within a period of less 
than five times the drug's half-life, whichever is longer 
• Use of any biologic within a period of 5 times its half-life 
• Use of vinegar or bleach baths within 2 weeks of starting the study 
 
 
7 
PARTICIPANT ENROLMENT 
 
7.1 
Consenting Participants 
Informed consent for participation in the study must be obtained before performing any study-specific 
procedures.  
 
Informed Consent Forms for enrolled participants and for participants who are not subsequently enrolled will be 
maintained at the study site in secure study files. Consent will be obtained by trained research study staff trained 
in taking informed consent. The study will be explained with the opportunity for the participant to ask 
questions. If a participant wishes to enter the study, a consent form will be completed and signed. 
 
7.2 
Screening for Eligibility 
After informed consent has been obtained, to determine participant eligibility for enrollment in the study, 
screening assessments will be performed within 1 week (-21 to -14 days) prior to starting the Washout period (-
14 to -1). All screening evaluations must be completed and reviewed to confirm that participants meet all 
eligibility criteria before Washout period and subsequent randomization on Day 1. Subjects will be asked to 
undergo a 2 week  (-14 to -1) washout period prior to the Baseline procedure. During the washout period, 
subjects will be asked to stop taking bleach or vinegar baths and start using cleanser and moisturizer provided 
by the sponsor on as needed bases.  

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At least 30 days of the participant’s medical history and medication use will be requested from their PCP or 
treating physicians and reviewed by the study staff. A complete physical examination including height, weight 
and vital signs will be performed.  
 
All screening assessments are listed in the Time and Events Table (Appendix A).  A participant must meet all 
inclusion criteria, and none of the exclusion criteria, to be enrolled and randomized in this study. The 
Investigator and team will maintain a screening log to record details of all persons screened and to confirm 
eligibility or record reasons for screening failure, as applicable. 
 
7.3 
Study Withdrawal and Withdrawal From Investigational Product and Stopping Criteria  
 
A participant may withdraw from the study at any time at his/her own request, or may be withdrawn at any time 
at the discretion of the Investigator. 
Reasons for withdrawal (participants who refuse to complete any remaining study visits) or discontinuation 
(participants who prematurely stop the application) at any time during the study may include, but are not 
limited, to the following: 
 
● For safety reasons, either at the discretion of the Investigator or at the participant’s request 
● For protocol violations at the discretion of AOBiome 
● Due to concomitant therapy that could interfere with the results of the study (the Investigator will report 
all such information on the CRFs and decide, in accordance with AOBiome, whether the participant is to 
be withdrawn).  
 
The reason for participant study withdrawal will be recorded in the electronic Case Report Form (eCRF). Data 
from participants withdrawing from the study will be considered evaluable up to the point at which they are 
withdrawn using the same criteria for evaluability as for participants who complete the study. 
 
7.4 
Screen Failures 
Data for screen and baseline failures, such as date and reason, will be collected in source documentation at the 
site 
and will be transmitted to AOBiome.  
 
7.5 
Early Termination 
Participants who have discontinued the study early will be evaluated by the Investigator at the Early 
Termination Visit.  See the list of assessments to be performed at the Early Termination Visit in the Time and 
Events Table (Appendix A). Participants with ongoing AEs or SAEs believed to be possibly related to 
investigational product (IP) will continue to be followed until resolution or for 30 days as warranted by the 
nature of the AE.  
 
8 
STUDY TREATMENT 
8.1 
Investigational Product 

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Under normal conditions of handling and administration, IP is not expected to pose 
significant safety risks to site staff. A Material Safety Data Sheet describing the 
occupational hazards and recommended handling precautions will be provided to site 
staff if required by local laws or will otherwise be available from the Sponsor upon request. 
 
Investigational product must be stored in a secure area under the appropriate physical 
conditions for the product. Access to and administration of the IP will be limited to the 
Investigator and authorized site staff. Investigational product must be dispensed or 
administered only to participants enrolled in the study and in accordance with the protocol. 
 
The Investigator is responsible for study treatment accountability, reconciliation, and record maintenance. The 
Investigator or designated site personnel must maintain study treatment accountability records throughout the 
course of the study. The responsible person(s) will document the amount of study treatment received from and 
returned to the sponsor and the amount administered to participants. The required accountability unit for this 
study will be the bottle. Discrepancies are to be reconciled or resolved.  
 
 
Product name: 
B244, 30ml/bottle 
Vehicle, 30ml/bottle 
Dosage form: 
B244 suspension 
Vehicle solution 
Unit dose strength: 
4x109 cfu/ml 
50nM Na2HPO4-2mM MgCl2 
(pH 7.6) 
Route/administration/duration: 
Topical application BID for 4 
weeks 
Topical application BID for 4 
weeks 
Dosing instruction: 
Subjects will apply a total of 8 
pumps of  IP per application 
to all affected areas twice-a-
day (i.e. 8 pumps in the 
morning and 8 pumps again at 
night). 
 
Applications should occur in 
the morning and at night for 4 
weeks. 
Subjects will apply a total of 8 
pumps of  IP per application 
to all affected areas twice-a-
day (i.e. 8 pumps in the 
morning and 8 pumps again at 
night). 
 
Applications should occur in 
the morning and at night for 4 
weeks. 
Physical description: 
Odorless, cloudy, light pink 
suspension 
Odorless, clear, and colorless 
suspension 
Manufacturer/source of 
procurement: 
AOBiome, LLC 
AOBiome, LLC 
 
The contents of the label will be in accordance with all applicable regulatory requirements. B244 and matching 
vehicle will be packaged in identical 30 ml white bottles.  
 
8.2 
Dose Changes  

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No dose changes are anticipated. 
 
8.3 
Storage conditions  
All investigational drug supplies in the study will be stored in a secure, refrigerated (2-8OC) safe place, under 
the responsibility of the Investigator or other authorized individual.  
 
8.4 
Description of Blinding Method 
This study will be double-blinded: neither Investigator(s), nor study participants, nor those involved in the 
conduct of the trial (including sponsor staff) will be aware of the treatment the participants are receiving. 
 
8.5 
Treatment Assignments: 
This is a double blind study. Participants will be assigned to study treatment in accordance with the 
randomization schedule generated for the allocation of vehicle or B244 prior to the initiation of the trial. 
Randomization will be centrally-based and performed using an appropriate IWRS (an automated randomization 
system).  
 
Each participant scheduled to receive investigational product (IP) will receive a randomization number at the 
time of randomization. The randomization number will be used to identify the study medication kit assigned to 
the participant and indicate the treatment to be administered to that participant.  
    
8.6 
Treatment Compliance 
Study treatment must be dispensed or administered according to procedures described 
herein. Only participants enrolled in the study may receive study treatment. Only authorized site personnel may 
supply study treatment. Participants will record use of the study medication utilizing the study diary at the time 
of use each day. Participants will review study medication compliance with the Investigator or designee. Any 
missed doses, timing, and reason for missed dose will be recorded in the eCRF. There should be no doubling of 
doses to make up for missed doses. If a dose is missed, the next dose of study medication should be taken as 
scheduled. 
 
Study staff will weigh the IP at the beginning of every visit.  Weight will be recorded in the eCRF. 
 
 
8.7 
Treatment Application 
Subjects will receive a kit containing study drug for application throughout the study. Subjects will be 
instructed in the use of the spray bottle and asked to self-administer the Investigational Product as follows: 
 
• Subjects will apply a total of 8 pumps of IP per application to all affected areas twice-a-day (i.e. 8 
pumps in the morning and 8 pumps again at night). 
• Subject should saturate the application area well. 
• Subjects may not wash the application site after applying the IP 

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• Subjects will be asked to let the product air dry. 
• While in use, one spray bottle may be stored at ambient temperature. The bottle that is not in current use 
by the subject, must be stored in the refrigerator.  DO NOT FREEZE. 
 
 
8.8 
Treatment of Investigational Product Overdose 
The sponsor does not recommend specific treatment for an overdose. Washing with conventional cleanser and 
water will remove the product. The Investigator will use clinical judgment to treat any overdose.  
 
8.9 
Product Accountability 
In accordance with federal and local regulatory requirements, the Investigator and designated site personnel 
must document the amount of investigational product dispensed to study participants, the amount returned by 
study participants, and amount received and returned to the sponsor, when applicable. Product accountability 
records must be maintained throughout the course of the trial. Any quality issue noticed with the receipt or use 
of an IP (deficient IP in condition, appearance, pertaining documentation, labeling, expiry date, etc.) should be 
promptly reported to the Sponsor, who will initiate a complaint procedure.  
All investigational product must be stored in a secure locked room with access limited to the Investigator and 
designated site personnel. Study product is to be stored in a refrigerator between 2-8 degrees C. Maintenance of 
a temperature log is required.  
Under no circumstances will the Investigator allow IP to be used other than as directed by this Clinical Trial 
Protocol, or dispose of IP in any other manner.  
 
8.10 Unblinding Procedures 
The Investigator may unblind a participant’s treatment assignment only in the case of emergency or in the event 
of a serious medical condition when knowledge of the study treatment is essential for the appropriate clinical 
management or welfare of the participant, as determined by the Investigator. It is preferred (but not required) 
that the Investigator first contacts the medical monitor to discuss options before unblinding the participant’s 
treatment assignment. The Investigator must notify the Sponsor as soon as possible when a participant’s 
treatment assignment is unblinded without revealing the treatment assignment of the unblinded participant 
unless that information is deemed important for the safety of participants currently in the study. The date and 
reason for the unblinding must be documented in the participant’s study record.  
The Medical Monitor may unblind the treatment assignment for any participant with an SAE. If the SAE 
requires that an expedited regulatory report be sent to one or more regulatory agencies, a copy of the report, 
identifying the participant’s treatment assignment, may be sent to clinical Investigators in accordance with local 
regulations and/or sponsor policy. 
 
8.11 Retrieval and Destruction of Investigational Product 
All partially used or unused treatments will be returned to the site as brought by study participants. A detailed 
IP log of the returned IP shall be established.  
The site will not destroy unused IP unless the Sponsor provides written authorization to the contrary. All used 
and unused bottles will be shipped to the authorized drug depot at the end of the study.  

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8.12 Permitted Medications 
All participants will be screened for concomitant medications prior to inclusion into the study.  
Any concomitant medication to treat adverse events will be recorded in the Concomitant Medication section of 
the eCRF. 
 
8.13 Prohibited Medications 
Participants will be prohibited from taking any of the following therapy, including topical corticosteroids, 
oral/IV/inhaled steroids, topical coal tar, topical sulfur, topical PDE-4 inhibitors, topical or oral antihistamines, 
topical antiseptics or antibiotics, topical antifungals, bleach baths, UVA or UVB phototherapy, oral 
antibiotics/antiviral/antifungal/antiseptic agents, systemic immunosuppressive or immunomodulatory agents, oral 
or topical probiotics for the duration of the trial.  
 
8.14 Rescue Medications 
Rescue medications will not be permissible in the trial should the study subject experience acute exacerbation of 
symptoms related to the treated disease. Subject may not start additional non-randomized medication and should 
immediately alert the study site PI. Subjects that need to start a rescue medication will be discontinued from the 
study and will be considered an Early Termination. 
 
8.15 Lifestyle Restrictions 
 
8.15.1 Use of cleanser and moisturizer during the trial  
 
At the beginning of the trial, subjects will be provided cleanser and moisturizer to use for the duration of the 
trial as needed. Subjects will be asked not to use any other cleansers, or moisturizers during the course of active 
treatment. (Baseline-Day 28). Although moisturizer is provided to subjects, its use it not mandatory in the study 
and may be used when and as needed only. If subjects do not have the need to use the moisturizer, they may 
forego its use all together.  
 
Although study cleanser is provided, subjects will be asked to refrain from washing the treatment areas with 
study cleanser starting from Washout through Day 28. Subjects may use cleanser to wash other parts of their 
body not treated during the study. At Day 28 (end of study drug application) subjects will be asked to return 
cleanser that were provided to them by the sponsor and will be asked to go back to their regular regimen and 
use their preferred cleanser. 
 
Provided moisturizer may be applied to the application site AFTER IP administration. Subject should apply the 
IP as instructed, let the site of application dry and only then apply the moisturizer.  
 
Subjects may not wash application area with cleanser and water AFTER each application.   
 
 

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8.16 Handling of Investigational Product 
 
Subjects will receive a kit containing two 30 ml white bottles. Each bottle will be used for one week and 
brought to all study appointments (Day 14, Day 28, Unanticipated visit, EOS visit).  Subjects will be asked to 
refrigerate bottles that are not in use. The bottle which is being used for treatment at a given week may be 
placed on the counter to be used during the treatment period. 
 
Subjects will be asked not to subject the treatment kit to conditions which are unnatural or harmful to the 
product, such as excessive heat (temperatures over 77°F (25°C) and freezing temperatures (at 0oC). Subjects 
may travel with their study medication but should not leave it in the hot car, outside in the cold temperatures 
etc. Subjects will also be asked not to tamper or cause damage to IP. 
 
 
8.17 Treatment compliance 
 
Each site participating in the trial will be instructed to assess subject’s compliance by weighing the 
investigational product and obtaining the weight of the bottle in grams pre and post application (after two weeks 
of use). Study personnel will be asked to take out bottles from the carton, weigh both bottle and record the 
weight. At the Day 14 visit, study personnel will need to weigh both bottles without the carton and record 
weight. This procedure should be followed every time study medication is dispensed and returned. Sites will be 
provided scales, which will be calibrated prior to each use. Study personnel will be instructed to record 
measurements into the eCRF.  
 
9 
ACTIGRAPHY WATCH 
At Baseline visit, subjects will be provided two actigraphy watches in order to accurately monitor subject’s 
sleep, activity and itching patterns. Subjects will be asked to wear one watch on each wrist 24 hours a day 
starting with the Baseline visit through Week 4 visit. Actigraphy watches are water resistant and may be left on 
when bathing/showering/swimming. However, for convenience, subjects may remove watches during the above 
mentioned procedures but will be asked to put watches back on once these procedures have been completed.  
 
Any deviations will be recorded in the CRF.   
10 CONTRACEPTION REQUIREMENTS 
 
Effective contraception is required for all women physiologically capable of becoming pregnant during study 
participation. Women of child-bearing potential must agree to use an acceptable form of contraception for up to 
2 weeks after the study completion as detailed. Women of childbearing potential are defined as any female who 
has experienced menarche and who is NOT permanently sterile or postmenopausal. Postmenopausal is defined 
as 12 consecutive months with no menses without an alternative medical cause. 
 
Effective contraception methods include:  
• Total abstinence (when this is in line with the preferred and usual lifestyle of the study participant). 
Periodic abstinence (i.e., calendar, ovulation, symptothermal, postovulation methods) and 

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withdrawal are not acceptable methods of contraception.  
• Use of oral, injected or implanted hormonal methods of contraception or placement of an IUD or 
IUS or other forms of hormonal contraception that have comparable efficacy, for example hormone 
vaginal ring or transdermal hormone contraception.  
• Use of barrier methods (i.e., condom, diaphragm) used with a spermicide (i.e., foam, cream, or gel 
that kills sperm) 
 
In case of use of oral contraception women should have been stable on the same pill for a minimum of 90 days 
before the baseline visit. 
Male participants of the study who are having sexual intercourse with a woman who can become pregnant must 
use an acceptable form of birth control while participating in the study. Additionally, male participants are 
expected to let their female partners know of their participation in a research study of a drug, and that the effects 
of the drug on an unborn baby and on a pregnant woman are unknown. Male participants will also be expected 
to provide their female partners with the contraception requirements information previously described and the 
study doctor’s contact information for questions.  
Payment for all aspects of obstetrical care, child-or related care will be the study participant’s responsibility.  
In case of pregnancy, Investigational Product should be discontinued and the Sponsor should be informed 
immediately. Follow-up of the pregnancy will be mandatory until the outcome is available. 
11 STUDY PROCEDURES 
11.1 Pre-screening procedures  
 
Study subjects will be recruited from among participating hospitals, clinics, and diagnostic centers or from 
general population under the responsibility of a participating Investigator. Prior to initiation of the recruitment 
phase, participating Investigators will identify a pool of potential study subjects. Each of these centers will 
identify potentially eligible patients in advance, by either reviewing past medical records and diagnoses, 
screening in clinics, referral from other physicians, or other sources of recruitment, to identify those aged 18 or 
older with clinical diagnosis of mild or moderate Atopic Dermatitis. Medical records from patient’s 
dermatologist or primary care physician to confirm the diagnosis are optional. Verbal confirmation of the 
diagnosis present for ³ 12 months is sufficient in order to fulfill this criterion.  
 
 
11.2 Informed Consent Procedures 
 
Eligible participants may only be included in the study after providing a consent using the IRB-approved 
informed consent. Informed consent must be obtained before conducting any study-specific procedures (i.e. all 
of the procedures described in the protocol). The process of obtaining informed consent should be documented 
in the participant’s source documents. The date of signing of informed consent (and withdrawal, if later 
withdrawn) should be documented in the CRF. 
 
11.3 Study Assessments 
 

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Study activities will take place according to the Time and Events table (Appendix A). 
 
11.4 Inclusion procedures  
 
Once all inclusion/exclusion criteria are fulfilled, the patient becomes eligible for randomization and inclusion 
into the treatment period. Treatment allocation will be performed as stated above in Section 8.5. Study 
medication will be delivered as stated in Section 8.7. Patients will be counseled on product application and 
diary completion.  
 
11.5 Timing of patient’s visits to the clinic 
Patients will be asked to report to the clinic for their scheduled appointments. If a subject is unable to schedule 
an appointment within the required time frame, study staff will be asked to reschedule the patient to a day when 
they are able to come in within predetermined time frame.  
 
 
11.6 Description by type of visit 
11.6.1 Screening Visit (-21 to -14) 
 
• 
informed consent completed and signed  
• 
inclusion and exclusion criteria  
• 
demographic data  
• 
medical/surgical history 
• 
perform Hanifin and Rajka criteria assessment  
• 
current medications 
• 
smoking status  
• 
physical exam 
• 
body weight 
• 
height measurement  
• 
obtain in clinic blood pressure and heart rate 
• 
urine pregnancy test (for women of childbearing potential) 
• 
Oral hygiene questions 
• 
EASI assessment 
• 
VAS assessment 
• 
Skindex 16 Questionnaire 
• 
IGA scoring assessment 
• 
blood for clinical chemistry, serology and hematology 
• 
Microbial content 
• 
provide subjects with study cleanser to start using for the Washout period 
• 
provide subjects with moisturizer to start using for the Washout period 
• 
Start AE monitoring 

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11.6.2 Washout Phase (-14 to -1) 
At the Screening visit once subjects become eligible for the study, they will be provided moisturizer and 
cleanser which need to be used for two weeks prior to the Baseline visit during the Washout phase of the study.  
Moisturizer use is not mandatory in the study and may be used when and as needed only. If subjects do not have 
the need to use the moisturizer, they may forego its use all together. However, subject will initiate the Washout 
phase once the chemistry and serology results become available from the lab. Once study staff receive those 
results, subjects will be notified by the telephone to start the Washout period phase and start using provided 
cleanser and moisturizer if needed.  Subject will be asked to stop using their regular cleanser and regular 
moisturizers for two weeks prior to their Baseline visit. Subjects will also be asked to stop using bleach or 
vinegar baths.  
 
• 
Call subjects to initiate Washout phase 
• 
provide counseling as needed 
• 
ask subjects to start using the provided cleanser 
• 
as subjects to start using the provided moisturizer 
• AE monitoring 
 
11.6.3 Study Day 1-Baseline visit (-1 to 0) 
• 
inclusion and exclusion criteria  
• 
perform Hanifin and Rajka criteria assessment  
• 
current medications 
• 
obtain in clinic BP and HR 
• 
EASI assessmentVAS assessment 
• 
Skindex 16 questionnaire 
• 
IGA scoring assessment 
• 
dispense actigraphy watches and educate subjects on its use 
• 
allocation of a randomized treatment kit number via IWRS 
• 
delivery of the corresponding pack of Investigational Product  
• 
obtain study medication weight for Investigational Product compliance 
• 
first application of Investigational Product (under medical supervision)  
• 
Microbial content 
• 
study counseling 
• 
study diary counseling 
• 
replenish cleanser and moisturizer when needed 
• 
obtain skin swabs 
• 
blood for biomarkers 
• 
AE monitoring 
 
11.6.4 Week 2, 4 (Day 14, 28) study visit 
• 
concomitant medications 
• 
body weight 

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• 
obtain in clinic heart rate and blood pressure 
• 
Oral hygiene questions 
• 
EASI assessment 
• 
VAS assessment 
• 
Skindex 16 Questionnaire 
• 
IGA scoring assessment 
• 
obtain study medication weight for Investigational Product compliance 
• 
collect study medication  
• 
dispense investigational product to patient (Week 2) 
• 
blood for biomarkers  
• 
blood for clinical chemistry and hematology 
• 
Microbial content (Week 4 visit only) 
• 
study diary 
• 
study counseling 
• 
provide subjects with study cleanser and moisturizer (when necessary only) 
• 
collect cleanser and moisturizer at Week 4 visit 
• 
collect actigraphy watches returned by the subject at Week 4 visit 
• 
recording of AEs if any 
• 
skin swabs (Week 4 only) 
 
 
11.6.5 Week 6 final visit (Day 42)-End of Study Visit 
• 
record concomitant medications  
• 
physical exam 
• 
body weight 
• 
obtain in clinic heart rate and blood pressure 
• 
Oral hygiene questions 
• 
EASI assessment 
• 
VAS assessment 
• 
Skindex 16 questionnaire 
• 
IGA scoring assessment 
• 
Skin swabs 
• 
blood for clinical chemistry and hematology 
• 
Blood for biomarkers 
• 
recording of AEs if any 
 
11.6.6  Unscheduled/Unanticipated Study visit 
If an event arises that requires patient to come in to the research center, subjects should be scheduled for the 
Unscheduled visit and assessments are performed based on investigator discretion. 
 
  

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11.6.7 Early Termination Visit 
Every attempt should be made to complete all visits during the defined window periods. Subjects who do not 
complete all required study visits and withdraw from the study before Week 6 final visit, will be asked to 
complete the Early Termination Visit. . 
 
During the visit, the following will be obtained: 
 
• 
record concomitant medications  
• 
physical exam 
• 
Body weight 
• 
obtain in clinic heart rate and blood pressure 
• 
Oral hygiene questions 
• 
EASI assessment 
• 
VAS assessment 
• 
Skindex 6 questionnaire 
• 
IGA scoring assessment 
• 
skin swab samples  
• 
obtain study medication weight for Investigational Product compliance ((if ET visit occurs before Week 
4 visit) 
• 
blood for biomarkers (if ET visit occurs before Week 4 visit) 
• 
blood for clinical chemistry and hematology (if ET visit occurs before Week 4 visit) 
• 
return Investigational product (if ET visit occurs before Week 4 visit) 
• 
return cleanser and moisturizer (if ET visit occurs before Week 4 visit) 
• 
return Actigraphy watch (if ET visit occurs before Week 4 visit) 
• 
return study diary (if ET visit occurs before Week 4 visit) 
• 
AE Monitoring 
 
 
12 METHODS OF ASSESSMENTS 
12.1 Skin Swab Samples 
 
Skin swab samples will be collected as Described in Appendix A. At Baseline, study staff will  
examine subject’s body surface area affected by AD and will be asked to identify only one area to be sampled. 
Once the area is identified, study staff will sample this particular treatment area. Sampling area should be noted 
in subject’s record, so that the same treatment area is then sampled at Day 28 and Day 42. Study staff will 
perform swabbing procedures during subject’s visit. Swab samples will need to be stored in the refrigerator 
until shipped at the end of the study to the SciSafe Laboratories. 
 
Swab samples may be subjected to DNA sequencing. Samples will be used for study of the skin microbial 
diversity and microbiome analysis While this is not our intention to study participants genome, participants 
sequence may be included as part of this analysis. Samples may be stored in the freezer until future research. 

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We will keep the specimen for up to 5 years, or until analyzed.  If we complete our research and no longer need 
to keep the specimens, we will destroy them. Specimens will be coded and no identifiable participant 
information will be used. Results will not be returned to the subjects or the investigators 
 
 
12.2 Blood Pressure Measurement 
 
Blood pressure readings will be obtained at every visit as described in Appendix A (schedule of events). Subject 
should be asked to remove all clothing that covers the location of cuff placement. The individual should be 
comfortably seated in the chair, with the legs uncrossed, and the back and arm supported, such that the middle 
of the cuff on the upper arm is at the level of the right atrium (the mid-point of the sternum). Neither the patient 
nor the observer should talk during the measurement. After 5 minutes sitting, serial clinic BP measurements and 
heart (x3) rate will be performed, using the non-dominant arm with a calibrated electronic sphygmomanometer. 
The average of the second and third readings will be documented. 
 
12.3 Physical Examinations  
 
The physical examination will be performed at Screening and at Unanticipated visit should one occur. The 
physical examination includes an assessment of general appearance and a review of systems (dermatologic, 
head, eyes, ears, nose, mouth/throat/neck, thyroid, lymph nodes, respiratory, cardiovascular, gastrointestinal, 
extremities, musculoskeletal, neurologic systems).  
 
12.4 Laboratory Assessments 
 
Blood samples should be taken using standard venipuncture techniques. Blood sampling will be performed 
according to the site SOPs. 
 
The following laboratory variables will be determined as outlined below: 
 
The following routine clinical chemistry, hematology and Lipid Panel will be performed  according to Time and 
Events table (Appendix A): Albumin, alkaline Phos, ALT, AST, Total Billirubin, BUN, BUN:Createnine ratio, 
Calcium, Chloride, Createnine, eGFR, Glucose, Potassium, Sodium, Uric Acid 
 
Lipid Panel: HDL Cholesterol, LDL cholesterol, Total Cholesterol, Triglycerides, VLDL Cholesterol, 
LDL/HDL Cholesterol Ratio, Non HDL Cholesterol. 
 
Angiotensin Converting Enzyme. 
 
Hematology: WBC, RBC, Hemoglobin, Hematocrit, Platelets, WBC Differential (Neutrophils, Lymphocytes, 
Monocytes, Eosinophils, Basophils), HbA1C 
 
Serology will only be done at Screening: HIV Ab, HCV Ab, HBsAg. 
 

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Patients will be asked to fast for at least 8 hours before all blood tests are done.  
 
The total blood volume collected for clinical labs for Screening visit will be approximately 20 ml of whole 
blood. Volume collected for subsequent visits would be approximately 10 ml of whole blood.  
 
Any value outside the normal range will be flagged for the attention of the Investigator or designee at the site. 
The Investigator or designee will indicate whether or not the value is of clinical significance. If the result of the 
clinical chemistry test from the samples taken during the screening phase is indicated as clinically significant, 
the study subject will NOT be allowed into the study.  
 
12.5 Oral Hygiene Questions 
 
Subjects will be asked questions regarding their oral hygiene and use of toothpaste. This will be will be 
administered at the times indicated in the Time and Events Table (Appendix A) 
12.6 EASI Assessment 
An EASI score is used to measure the extent (area) and severity of atopic eczema (Eczema Area 
and Severity Index). The score will be calculated at  the times indicated in the Time and Events Table 
(Appendix A).The investigator will assess improvement of eczema based on intensity and severity of the 
disease. EASI score sheet can be found in Appendix C. 
 
The severity strata for the EASI are as follows:  
0 = clear 
0·1-1·0 = almost clear 
1·1-7·0 = mild 
7·1-21·0 = moderate 
21·1-50·0 = severe 
50·1-72·0 = very severe  
 
 
12.7 Subject reported outcomes/Skindex16  
Skindex-16 instrument will be administered at the times indicated in the Time and Events Table (Appendix A). 
The participant will answer a questionnaire examining the relationship between the patient’s skin health and 
quality of life.  
 
Skindex16 questionnaire can be found in Appendix D. 
 
12.8 Hanifin and Rajka Diagnostic Criteria for Atopic Dermatitis (AD) 
At Screening and Baseline, all subjects must have atopic dermatitis, as defined by the Hanifin and Rajka 
criteria, which involves a minimum of 10% and a maximum of 30% body surface area. Hanifin and Rajka 
criteria can be found in Appendix E  
Major criteria: Must have three or more of: 
 

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1. Pruritus 
2. Typical morphology and distribution 
• 
Flexural lichenification or linearity, erythema, scaling, serum-crust 
3. Chronic or chronically-relapsing dermatitis 
4. Personal or family history of atopy (asthma, allergic rhinitis, atopic dermatitis) 
 
Minor criteria: Should have three or more of: 
1. Xerosis 
2. Ichthyosis, palmar hyperlinearity, or keratosis pilaris 
3. Immediate (type 1) skin-test reactivity 
4. Raised serum IgE 
5. Early age of onset 
6. Tendency toward cutaneous infections (especially S aureus and herpes simplex) or impaired cell-
mediated immunity 
7. Tendency toward non-specific hand or foot dermatitis 
8. Nipple eczema 
9. Cheilitis 
10. Recurrent conjunctivitis 
11. Dennie-Morgan infraorbital fold 
12. Keratoconus 
13. Anterior subcapsular cataracts 
14. Orbital darkening 
15. Facial pallor or facial erythema 
16. Pityriasis alba 
17. Anterior neck folds 
18. Itch when sweating 
19. Intolerance to wool and lipid solvents 
20. Perifollicular accentuation 
21. Food intolerance 
22. Course influenced by environmental or emotional factors 
23. White dermographism or delayed blanch 
 
 
12.9 Visual Analog Scale (VAS) 
VAS will be performed as a measure of pruritus at the times indicated in the Time and Events Table (Appendix 
A). Subjects will be asked to report their pruritus symptoms over the 24 hrs period before the visit.  
 
VAS score can be found in Appendix F.  
 
The following VAS categories will be used:  
0=no pruritus 
>0–<4 points=mild pruritus 
≥4–<7 points=moderate pruritus 

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≥7–<9 points=severe pruritus 
≥9 points=very severe pruritus 
 
 
 
 
 
 
12.10 Investigator Global Assessment Score (IGA) 
 
IGA will be performed at the times indicated (Screening, Baseline, Weeks 2, 4, and 6) in the Time and Events 
Table of the Protocol. The Investigator will assess improvement of Atopic Dermatitis based on the 5-point 
severity scale summarized below.  
 
IGA score can be found in Appendix G.   
 
 
 
12.11 Actigraphy Watch 
Subjects will be asked to wear two Actiwatch Spectrum watches on each wrist  for the duration of the study, 
starting with the Baseline visit. The device is the size of a wrist watch, is water resistant, and has a plastic wrist 
band that is hypoallergenic so it is not likely to cause any skin problems. The device has been used by 
thousands of research subjects in hundreds of clinical trial with no significant problems for the subjects.  
Two watches will be given to subjects at Baseline visit.  Subjects will be asked to put both watches on their 
wrists during the visit and wear both watches at all times until Day 28 visit. Subjects will be allowed to take 
watches off for bathing, swimming and/or work or other activities prohibiting wear of such equipment. Study 
site personnel will go over the procedures with each patient making sure patient understand the procedures.  
The actigraphy data will be used to determine the estimates of total sleep time (TST) and total wake time 
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Upon the start of the study, Philips will provide each site with training and provide each site with the number of 
devices it needs, monitor the battery status of each device and replace as needed, and provide a 24/7 help desk.  
 
12.12 Biomarkers 
In addition to the blood drawn for the safety laboratory assessments, additional blood will be collected for the 
biomarker analysis at Day 1B (Baseline), Day 14, Day 28 and Day 42 
Subjects must be fasting for at least 8 hours Samples will be processed on site and stored in the - 80○ C 
Approximately 20 ml of whole blood will be drawn for biomarkers at each visit. Patients will be asked to fast 
for at least 8 hrs before blood for biomarkers is drawn.  
 
Blood samples for biomarkers will be taken according to Appendix A and processed according to the lab 
manual. Biomarkers will be evaluated for cytokines, chemokines, inflammatory markers and immune response.  
 
 
12.13 Microbial Content 
 
Microbial content samples will be taken from one affected area and analyzed for microbial levels of Staph. 
aureus. Cup scrub technique will be used in recovery of microorganisms from the skin.  
 
At Screening, study staff will examine subject’s body surface area affected by AD and will be asked to identify 
only one area to be sampled. Once the area is identified, study staff will sample this particular treatment area. 
Sampling area should be noted in subject’s record, so that the same treatment area is then sampled at Baseline 
and Week 4 visits.  
 
Skin sampling will be conducted by the trained study staff in the study office.   
Microorganisms are recovered from the site by pressing a cylinder against the affected skin with sufficient 
pressure to form a seal and instilling recovery buffer (phosphate buffer) into the cylinder. The surface of the 
skin is then mechanically ‘scrubbed’ with a glass rod for 2 minutes. The fluid is pipetted from the cylinder into 
a test tube for further analysis.  
 
Sterile supplies will be provided by the Sponsor.   
 
Samples may be stored frozen for up to 5 years for verification of results and possible characterization of strains 
recovered from subjects.  
 
 
12.14 Sample Shipment 
All clinical chemistry samples are to be shipped to the central laboratory where samples will be analyzed. 
Please refer to the Lab Manual for sample shipping temperature and Lab destination information.  
 
 

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13  SAFETY ASSESSMENTS 
13.1 Compliance 
 
Participants will be asked to bring study medication with them to each scheduled visit. Study site will be 
provided with a scale and weight of the study medication will be obtained before the first use and at each visit.   
 
13.2 Pregnancy Reporting 
 
Any pregnancy will be reported by study participants during their study participation. Participants who report 
pregnancy or lactation during the review of inclusion/exclusion criteria prior to randomization will not be 
enrolled in the trial. In case of pregnancy, Investigational Product should be discontinued and the Sponsor 
informed. Follow-up of the pregnancy will be mandatory until the outcome is available.  
13.3 Study Completion 
 
A completed participant is one who has completed all study visits. Day 42 study visit is defined as the 
participant’s last visit. 
 
13.4 Subject Withdrawal Criteria 
A subject may withdraw from the study at any time at his/her own request, or may be withdrawn at any time at 
the discretion of the Investigator for safety, behavioral, or administrative reasons.  
Reasons for withdrawal (subjects who refuse to return for any remaining study visits) or discontinuation 
(subjects who prematurely stop the application) at any time during the study may include, but are not limited, to 
the following: 
 
● For safety reasons, either at the discretion of the Investigator or at the subject’s request 
● For protocol violations at the discretion of AOBiome 
● Due to concomitant therapy that could interfere with the results of the study (the Investigator will report 
all such information on the CRFs and decide, in accordance with AOBiome, whether the subject is to be 
withdrawn).  
All premature discontinuations and their causes must be carefully documented by the Investigator on the CRF 
or if needed on the AE form.   
If, for any reason, a subject is withdrawn before completing the final visit, the reason for termination will be 
entered on the CRF.  All data gathered on the subject prior to termination will be made available to AOBiome. 
Subjects not completing the entire study should be fully evaluated when possible.  The appropriate CRFs should 
be completed. 
 
If the subject chooses to withdraw before completing the study, the subject should notify the study coordinator 
who will instruct the subject on completion of assessments for End of Study (EOS) visit  Appendix A.  For 

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subjects who refuse to complete the assessments for their early termination, every attempt must be made to 
check on their status, using any mode of communication such as telephone, email, fax, or text. 
 
 
14 EFFICACY ASSESSMENTS 
Efficacy endpoints will be descriptively summarized and will include the number of observations, mean, 
median, standard deviation, minimum, and maximum of scores/values at all applicable time points and for all 
treatments in the ITT Population. 
EASI score at Baseline, Day 14, 28 and 42 will be compared to the baseline score and will be descriptively 
summarized as mean, median, standard deviation, minimum, and maximum of scores/values at all applicable 
time points and for all treatment groups. 
15 STATISTICAL CONSIDERATIONS 
15.1 Sample Size 
A total sample size of 130 randomized subjects, allows for a dropout rate of 20%. 
  
A sample size of 52 per group achieves >80% power to reject the null hypothesis of equal means when the 
population mean difference is μ1 – μ2 = (-1.4) – (-5.3) = 3.9 with a standard deviation for both groups 
of 7.0 and with a significance level (alpha) of 0.025 using a one-sided two-sample equal-variance t-test. 
  
The power calculation assumes baseline treatment group means of 14 in EASI score. The change from baseline 
to Day 28 is assumed to be -1.4 for the vehicle group and -5.3 for the active group; this represents about 
38% reduction in EASI score for the active group and 10% reduction in EASI score for the placebo group. 
 
15.2 Populations for Analysis 
ITT: includes all randomized participants.   
Safety: includes all subjects who received at least 1 dose of study medication.  
Per Protocol: subjects who administered at least 50 % of IP, have at least one baseline and post baseline in 
clinic visit and did not have any major protocol violations.  
15.3 Data Analysis 
The analyses will be conducted on all participant data when the trial ends.  Data will be presented by treatment 
group and overall.   
 
Categorical variables will be summarized by frequency distributions (number and percentages of subjects) and 
continuous variables will be summarized by descriptive statistics (mean, standard deviation etc). 
Adverse events will be summarized by treatment group using the Medical Dictionary for Regulatory Activities 
(MedDRA) system organ class and preferred term. Separate tabulations will be produced for all treatment-
emergent AEs (TEAEs), treatment-related AEs (those considered by the Investigator as at least possibly drug 

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related), SAEs, discontinuations due to AEs, and AEs ≥Grade 3 severity. By-subject listings will be provided 
for any deaths, SAEs, and AEs leading to discontinuation. 
Descriptive statistics for each treatment group will be provided for clinical laboratory values (e.g., hematology, 
serum chemistry, and urinalysis) and vital signs data, presented as both actual values and changes from baseline 
relative to each on-study evaluation. Abnormal clinical laboratory values will be listed.  
All data will be provided in by-subject listings. 
 
15.3.1 Disposition 
A tabulation of the disposition of subjects will be presented, including the number enrolled, the number 
randomized, the number treated, and the reasons for study discontinuation. Summaries of the number in each 
analysis set will be presented. Entry criteria violations and protocol deviations will be listed. 
 
15.3.2 Demographic and Baseline 
Demographic and baseline characteristic data summarization will be performed in order to descriptively assess 
the comparability of treatment groups. Data to be tabulated will include age, race, ethnicity, height, weight, and 
BMI,  as well as baseline characteristics related to medical history. 
 
15.4 Safety Analyses 
15.4.1 Definitions  
All adverse events recorded during the study will be coded according to MedDRA.  
15.4.2 Adverse Events  
All adverse events (AEs) recorded during the study through the date of randomization through 28 days after the 
last dose of study drug will analyzed.  
All AE’s will be coded according to MEDRA and summarized using SOC and Preferred term.  
 
AE’s will be summarized using incidence rates. Therefore, each subject will only contribute once for a given 
adverse event SOC or PT.  
 
A summary of the incidence of any adverse event, SAE, and adverse events leading to discontinuation will be 
presented.  Summaries will display, by treatment group, the incidence of patients with events, the frequency of 
patients with events within each primary system organ class and by preferred terms. For each preferred term and 
each system organ class a patient will be counted only once. For summaries on severe or drug-related AE, for a 
given patient, the highest severity or relationship for a specific preferred term will be considered.  
 
15.4.3 Deaths and Serious Adverse Events  
Serious adverse events and events leading to death will be summarized overall and by primary system organ 
class and preferred term.  
 
15.4.4 Adverse Events leading to treatment discontinuation  
Adverse events leading to treatment discontinuation will be summarized overall and by primary system organ 
class and preferred term.  
 

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15.4.5 Efficacy Analyses 
All efficacy analyses will be performed on the ITT population and PP population. P-values from analyses of 
efficacy endpoints must be interpreted in an exploratory fashion since there is no adjustment for multiple 
endpoints in this study. 
 
The EASI score and other continuous efficacy endpoints will be summarized using descriptive statistics at 
Baseline, Day 14, Day 28 and Day 42 for actual values and change-from-baseline values. The difference in 
treatment groups in change-from-baseline values at post-baseline visits will be analyzed using a mixed Analysis 
of Variance with one between-subject variable (active group vs. vehicle group) and one within-subject variable 
(Baseline vs. Day 14, Day 28, Day 42). The interaction between within-subject and between-subject variable 
will be included in the mixed model to test if any difference in treatment groups at post-baseline visits. Outputs 
from the analysis will include the p-value for the treatment group and interaction factor separately. The p-value 
should be evaluated at a significance level of 0.05. 
 
Additionally, a one-sided two-sample equal-variance t-test will be performed as a supportive test to analyze the 
differences in treatment group means in change from baseline at each post-baseline visit (Day 14, Day 28, Day 
42). Output from the analysis will include a 95% confidence interval for the difference in treatment groups 
means at each visit, and the p-values from the hypothesis test of no difference in treatment group means at each 
visit. The p-value should be evaluated at a one-sided significance level of 0.025. 
 
 
15.4.6 Handling of dropouts or missing data  
Missing data will not be imputed for analysis.  
Subjects who dropout after enrollment but prior to randomization will be replaced.  
 
15.5 Clinical Trial Protocol deviations  
At minimum, the following deviations will be summarized on the ITT patient population:  
 
• 
Inclusion or exclusion criteria not satisfied.  
• 
Deviations related to the Investigational Product administration  
• 
Not permitted concomitant medications.  
16 ADVERSE EVENTS (AE) AND SERIOUS ADVERSE EVENTS (SAE) 
The Investigator and study staff are responsible for detecting and recording AEs and SAEs during scheduled 
safety evaluations and whenever such information is brought to their attention. This section of the protocol 
provides definitions and detailed procedures to be followed. 
 
16.1 Definition of an AE 
An AE is any untoward medical occurrence in a study participant which is temporally associated with the use of 
a medicinal product, regardless of its potential relationship to the medicinal product.  An AE, therefore, can be 
any unfavorable or unintended sign, including an abnormal symptom, or disease (new or exacerbated), whether 
or not related to the investigational product (IP).   

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Examples of an AE include: 
● Exacerbation of a pre-existing condition including either an increase in frequency and/or intensity of the 
condition. 
● New conditions detected or diagnosed after investigational product administration even though it may have 
been present prior to the start of the study. 
● Signs, symptoms of a drug interaction. 
● Signs, symptoms of a suspected overdose of either investigational product or a concurrent medication 
(overdose per se should not be reported as an AE/SAE). 
 
AEs may include pre- or post-treatment events that occur as a result of protocol-mandated procedures (i.e., 
modification of participant’s previous therapeutic regimen). 
 
16.2 Definition of a SAE 
A serious adverse event is any untoward medical occurrence that, at any dose: 
 
a) results in death. 
b) is life-threatening. 
NOTE:  The term 'life-threatening' in the definition of 'serious' refers to an event in which the participant was at 
risk of death at the time of the event.  It does not refer to an event, which hypothetically might have caused 
death, if it were more severe. 
c) requires hospitalization or prolongation of existing hospitalization. 
NOTE:  In general, hospitalization signifies that the participant has been detained (usually involving at least an 
overnight stay) at the hospital or emergency ward for observation and/or treatment that would not have been 
appropriate in the physician’s office or out -patient setting.  Complications that occur during hospitalization are 
AEs.  If a complication prolongs hospitalization or fulfills any other serious criteria, the event is serious.  When 
in doubt as to whether “hospitalization” occurred or was necessary, the AE should be considered serious. 
Routine hospitalizations or elective surgeries are generally not regarded as SAEs.  
 
d) results in disability/incapacity 
NOTE:  The term disability means a substantial disruption of a person’s ability to conduct normal life functions.  
This definition is not intended to include experiences of relatively minor medical significance such as 
uncomplicated headache, nausea, vomiting, diarrhea, influenza, and accidental trauma (e.g. sprained ankle) 
which may interfere or prevent everyday life functions but do not constitute a substantial disruption. 
e) is a congenital anomaly/birth defect 
f) Medical or scientific judgment should be exercised in deciding whether reporting is appropriate in other 
situations, such as important medical events that may not be immediately life-threatening or result in 
death or hospitalization but may jeopardize the participant or may require medical or surgical 
intervention to prevent one of the other outcomes listed in the above definition.  These should also be 
considered serious.  Examples of such events are invasive or malignant cancers, intensive treatment in 

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an emergency room or at home for allergic bronchospasm, blood dyscrasias or convulsions that do not 
result in hospitalization, or development of drug dependency or drug abuse. 
16.3 Time Period, Frequency, and Method of Detecting AEs and SAEs 
 
All AEs occurring after administration of the first dose of study medication and on or before the final 
assessment must be reported as AEs.  All AEs must be recorded irrespective of whether they are considered 
drug-related. 
At each assessment in the period defined above, AEs will be evaluated by the Investigator and recorded. 
Any AEs already documented at a previous assessment and designated as ongoing, should be reviewed at 
subsequent assessments as necessary.  If these have resolved, this should be documented.  Changes in intensity 
or frequency of AEs should be recorded as separate events (i.e., a new record started). 
The recording of AEs and SAEs are described in Section 164 (“Recording of AEs and SAEs”). 
 
16.4 Recording of AEs and SAEs 
All clinical events, including either observed or volunteered problems, complaints or symptoms are to be 
recorded on the Adverse Events page(s) of the CRF.  The need to capture this information is not dependent 
upon whether the clinical event is associated with study treatment. Adverse clinical events resulting from 
concurrent illnesses or reactions to concurrent medications are also to be recorded.  In order to avoid vague, 
ambiguous, or colloquial expressions, the AE should be recorded in standard medical terminology rather than 
the participant’s own words. 
Each adverse clinical event is to be evaluated for duration, intensity, and whether the event may be associated 
with the investigational product (IP) or other causes.  Start and stop dates, relationship to investigational product 
(IP), medical management, and alternative causality of event must be recorded in the Adverse Events section of 
the CRF.  AEs believed to be possibly related to investigational product (IP) must be followed until resolution. 
 
16.5 Evaluating AEs and SAEs 
 
16.5.1 Severity Rating 
The severity of an adverse event (AE and SAE) is to be scored according to the following scale: 
Mild 
Awareness of sign or symptom, but easily tolerated 
Moderate 
Discomfort enough to cause interference with usual 
activity 
Severe 
Incapacitating with inability to work or perform usual 
activity 
An AE that is assessed as severe should not be confused with a SAE.  An event is defined as ‘serious’ when it 
meets one of the pre-defined outcomes as described in Section16.2  “Definition of a SAE”. 
 
16.5.2 Relationship to Investigational product (IP)  
SAEs will be classified as “not related” or “related” (including unknown).  

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For AEs, the relationship to study treatment is to be assessed according to the following definitions: 
Definitely not related: The AE is definitely not related to the drug.  This designation should be reserved 
for those events which occur prior to study treatment or for those events which cannot be even remotely 
related to study participation (e.g., injuries sustained in an automobile accident). 
 
Unlikely related: There is no reasonable association between the study treatment and the suspected 
event and the event could have been produced by the participant 's clinical state or other modes of 
therapy administered to the participant. 
 
Possibly related:  The suspected adverse event may or may not follow a reasonable temporal sequence 
from study treatment administration but seems to be the type of reaction that cannot be dismissed as 
unlikely.  The event could have been produced or mimicked by the participant 's clinical state or by 
other modes of therapy concomitantly administered to the participant. 
 
Probably related: The suspected adverse event follows a reasonable temporal sequence from study 
treatment administration, abates upon discontinuation of the treatment, and cannot be reasonably 
explained by the known characteristics of the participant 's clinical state. 
 
Definitely related: This designation should be reserved for those events which have no uncertainty in 
their relationship to treatment administration. 
 
 
 
 
16.6 Pregnancy 
 
Any pregnancy that occurs in a female participating in the study must be reported to the Sponsor within 24 hrs 
of learning of the pregnancy. Follow-up must occur to determine the outcome of the pregnancy (including 
premature termination) and the status of mother and child.  Pregnancy complications and elective terminations 
for medical reasons must be reported as AEs or SAEs. Spontaneous abortions must be reported as an SAE. 
Any SAE occurring in association with a pregnancy and considered by the Investigator as possibly related or 
related to the investigational product must be promptly reported to the Sponsor, even if the event occurred after 
the participant completed the study. 
The Investigator must attempt to collect pregnancy information on any female partners of male participants who 
become pregnant while the male participant is enrolled in the study. Pregnancy information must be reported to 
the Sponsor as described above. 
 
16.7 Prompt Reporting of SAEs to the Sponsor 
 
In the case of a Serious Adverse Event the Investigator must immediately: 
 
• 
SEND (within 1 working day, by email) the signed and dated corresponding page(s) in the Case Report 
Form to the representative of the Monitoring Team whose name, address and fax number appear on the 

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Clinical Trial Protocol, or to a designated Safety fax number provided by the Monitoring Team, as well 
as to the Central Database number;  
• 
ATTACH a photocopy of all examinations carried out and the dates on which these examinations were 
performed. Care should be taken to ensure that the patient's identity is protected and the patient's 
identifiers in the Clinical Trial are properly documented on all copies of source documents provided to 
the Sponsor. For laboratory results, include the laboratory normal ranges;  
• 
Follow-up of any Serious Adverse Event that is fatal or life threatening should be provided within one 
additional calendar week. The treatment code will be unblinded for reporting of Serious Adverse Events 
that are unexpected and reasonably associated with the use of the Investigational Product.  
Follow-up  
• 
The Investigator should take all appropriate measures to ensure the safety of the patients, including 
referral to a specialist if indicated. Notably he/she should follow up the outcome of any adverse events 
(clinical signs, laboratory values or other, etc) until the return to normal or stabilization of the patient's 
condition;  
• 
In the case of any serious adverse event, the patient must be followed up until clinical recovery is 
complete and laboratory results have returned to normal, or until progression has been stabilized. This 
implies that follow-up may continue after the patient has left the Clinical Trial and that additional 
investigations may be requested by the Monitoring Team;  
• 
In case of any serious adverse event brought to the attention of the Investigator at any time after 
cessation of Investigational Product and considered by him/her to be caused by the Investigational 
Product with a reasonable possibility, this should be reported to the Monitoring Team.  
 
. 
 
 
AOBiome Reportable Events Hotline 
Email: safety@aobiome.com 
 
 
17 ETHICAL AND REGULATORY STANDARDS 
17.1 Ethical Conduct of Study 
This clinical trial was designed and shall be implemented, executed, and reported in accordance with the ICH 
Harmonized Tripartite Guidelines for Good Clinical Practice, with applicable local regulations (including US 
Code of Federal Regulations Title 21), and with the ethical principles laid down in the Declaration of Helsinki. 
The Investigator(s) should conduct the study in accordance with this protocol, the Declaration of Helsinki and 
ICH GCP guidelines and FDA regulations.  The Investigator(s) and the Sponsor will sign the protocol and study 
contract, to confirm agreement.  The Investigator(s) will not implement any amendment (deviation or changes 
of the protocol) without agreement by the Sponsor and IRB approval, except where necessary to eliminate 

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immediate hazard(s) to study participants, or when change(s) involve only logistical or administrative aspects of 
the study. 
Records that may reveal the identities of participants must be well protected, with consideration given to 
confidentiality and the right to privacy of participants. 
17.2 Laws and Regulations 
This Clinical Trial will be conducted in compliance with all international laws and regulations, and national 
laws and regulations of the countries in which the clinical trial is performed, as well as any applicable 
guidelines. The trial will be registered and updated on www.clintrials.gov and on other sites, as deemed 
appropriate.  
17.3 Informed Consent 
Each participant must be provided with a statement that the investigation involves research and that the IRB has 
approved solicitation of participants to participate; a fair explanation of the procedures to be followed and their 
purposes, including identification of any procedures which are experimental; a description in lay language of 
any possible side effects; a description of any attendant discomforts and risks reasonably to be expected; a 
description of any benefits reasonably to be expected; a disclosure of any appropriate alternative procedures that 
might be advantageous for the participant; an offer to answer any inquiries concerning the procedures, and 
instruction that the person is free to withdraw consent and discontinue participation in the project or activity at 
any time without prejudice to the participant.  Payment to research participants for taking part in the study is 
based on time and inconvenience.   All information concerning payment, including the schedule of payments, 
must be set forth in the informed consent, including a disclosure that the Investigator is being paid to perform 
the stated research. 
A participant must give consent to take part in the study.  Participants below the age of majority in the 
municipality must give written assent to participate in this study.  This consent must be dated and retained by 
the Principal Investigator as part of the study records.  A downloadable digital copy shall be given to the person 
signing the form.  The informed consent process must be documented in the participant’s source documents. 
The Investigator agrees that the Sponsor, its employees or agents will have the right from time to time during 
the course of this study to audit and review pertinent medical records relating to this clinical trial.  A statement 
will be obtained from each person participating in the study permitting the release of his/her medical records as 
necessary for inspection by authorized personnel of the Sponsor, FDA, and the staff managing the clinical 
study.  
The release of medical records and the review of the contents will be in compliance with the Health Insurance 
Portability and Accountability Act of 1996 (HIPAA). 
 
17.4 Institutional Review Board/Independent Ethics Committee (IRB/EC) 
 
The protocol and informed consent form and the electronic version of the consent for this study must be 
approved by the IRB.  A copy of the Letter of Approval from the Board, which contains specific identification 
of the documents approved, must be received by the Sponsor prior to shipment of drug supplies to the Principal 
Investigator. 
All changes to the protocol, as well as a change of Investigator, must also be approved by the IRB and 
documentation of this approval provided to the study monitor.  Records of the IRB’s review and approval of all 
documents pertaining to the study must be kept on file by the Principal Investigator and are subject to FDA 

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inspection at any time.  IRB renewal for approval is required each year.  The Investigator is to inform 
AOBiome, in writing, of the approval to continue the study.   
 
17.5 Clinical Monitoring/Record Keeping 
There shall be no alterations in the protocol design without the written consent and approval of the Sponsor and 
the approval of the IRB, except in the case that participants are at immediate risk without immediate 
implementation of such alterations.  In the aforementioned situation, the site should notify the Sponsor and IRB 
of the deviation as soon as possible, and should seek the written consent and approval of the Sponsor and the 
approval of the IRB. 
All results of this trial must be recorded on eCRFs.  Each participant who has been randomized must have a 
completed eCRF.  Reasons for termination must be stated in the early termination section.  Study participants 
are not to be identified by name on eCRFs, but rather by coded identifiers and participant initials.   
The study monitor will verify the accuracy of the data by reviewing pertinent source documents such as office 
records or hospital charts of the participants. 
Study records include eCRFs, signed FDA Form 1572, original reports of test results, and signed electronic 
informed consent forms.  IRB approval letters and other documents pertaining to the conduct of the study are to 
be kept on file by the Investigator.   If the study files are assigned to someone else or removed to another 
location, the Investigator is to notify the study monitor or Sponsor in writing of the change.  All study records 
are subject to FDA inspection at any time. 
All information supplied to the Investigator by the Sponsor before, during, and after the study is confidential.  
Such information is to be used solely in connection with the clinical study.  The study protocol, IB, and any 
other pertinent study related materials or records provided are to be maintained in a confidential manner, 
reviewed carefully with attention to admonitions and returned to the Sponsor upon request.  No part of these 
materials may be reproduced or transmitted in any form without prior written permission from the Sponsor. 
18  ADMINISTRATIVE RULES 
18.1 Curriculum Vitae 
An updated, signed, and dated copy of the curriculum vitae with the experience, qualification and training for 
each Investigator and/or Sub-Investigator(s) will be provided to the Sponsor prior to the beginning of the 
Clinical Trial.  
18.2 Archiving of Study Documentation 
The Investigator must maintain confidential all study documentation, and take measures to prevent accidental or 
premature destruction of these documents.  
It is recommended that the Investigator retain the study documents at least fifteen (15) years after the 
completion or discontinuation of the Clinical Trial. However, applicable regulatory requirements should be 
taken into account in the event that a longer period is required.  
The Investigator must notify the Sponsor prior to destroying any study essential documents within the fifteen 
(15) year period following the Clinical Trial completion or discontinuation.  
If the Investigator's personal situation is such that archiving can no longer be ensured by him/her, the 
Investigator shall inform the Sponsor and the relevant records shall be transferred to a mutually agreed upon 
designee.  
 
18.3 Internal Safety Review Committee 

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An internal safety review committee will be set up to protect the ethical and safety interests of participants and 
to protect the scientific validity of the study. Adhoc safety interim analyses might be performed by an 
independent statistician if the safety review committee identifies potential safety signals during its routine 
blinded safety review. The details for the analysis plan will be documented in the trial’s Statistical Analysis 
Plan. 
  
19 STUDY MONITORING 
19.1 Responsibilities of the Investigator(s)  
The Investigator(s) undertake(s) to perform the Clinical Trial in accordance with this Clinical Trial Protocol, 
ICH guidelines for Good Clinical Practice and the applicable regulatory requirements.  
 
The Investigator is required to ensure compliance with all procedures required by the Clinical Trial Protocol and 
by study procedures provided by the Sponsor (including security rules). The Investigator agrees to provide 
reliable data and all information requested by the Clinical Trial Protocol (with the help of the Case Report Form 
[CRF], Discrepancy Resolution Form [DRF] or other appropriate instrument) in an accurate and legible manner 
according to the instructions provided and to ensure direct access to source documents to Sponsor 
representatives. If any particular circuits have to be defined (e.g., e-CRF, Fax), particular attention should be 
paid to the confidentiality of the patient's data to be transferred. The Investigator may appoint such other 
individuals as he/she may deem appropriate as Sub-Investigators to assist in the conduct of the Clinical Trial in 
accordance with the Clinical Trial Protocol. All Sub-Investigators shall be timely appointed and listed. The Sub-
Investigators will be supervised by and under the responsibility of the Investigator. The Investigator will 
provide them with a Clinical Trial Protocol and all necessary information.  
 
19.2 Responsibilities of the Sponsor  
 
The Sponsor of this Clinical Trial is responsible to Health Authorities for taking all reasonable steps to ensure 
the proper conduct of the Clinical Trial Protocol as regards ethics, Clinical Trial Protocol compliance, integrity 
and validity of the data recorded on the Case Report Forms. Thus, the main duty of the Monitoring Team is to 
help the Investigator and the Sponsor maintain a high level of ethical, scientific, technical and regulatory quality 
in all aspects of the Clinical Trial. At regular intervals during the Clinical Trial, the site will be contacted, 
through monitoring visits, letters or telephone calls, by a representative of the Monitoring Team to review study 
progress, Investigator and patient compliance with Clinical Trial Protocol requirements, and any emergent 
problems. During these monitoring visits, the following but not exhaustive list of points will be scrutinized with 
the Investigator: patient informed consent, patient recruitment and follow-up, Serious Adverse Event 
documentation and reporting, outcome events documentation and reporting, Investigational Product allocation, 
patient compliance with the Investigational Product regimen, Investigational Product accountability, 
concomitant therapy use and quality of data.  
 
19.3 Source document requirements  
According to the ICH guidelines for Good Clinical Practice, the Monitoring Team must check the Case Report 
Form entries against the source documents. The Informed Consent Form will include a statement by which the 
patient allows the Sponsor’s duly authorized personnel, the Ethics Committee (IRB/EC), and the regulatory 

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authorities to have direct access to source data which supports the data on the Case Report Forms (e.g., patient's 
medical file, appointment books, original laboratory records, etc.). These personnel, bound by professional 
secrecy, must keep confidential all personal identity or personal medical information (according to 
confidentiality rules).  
 
19.4 Use and completion of Case Report Forms (CRFs) and additional requests  
The sponsor or CRO will be responsible for activities associated with the data management 
of this study. This will include setting up a relevant database and data transfer mechanisms, 
along with appropriate validation of data and resolution of queries. Data generated within this 
clinical study will be handled according to the relevant SOPs of the data management and 
biostatistics departments of sponsor or CRO. 
  
For Electronic Data Capture (EDC): 
Study sites will enter data directly into an EDC system by completing the eCRF via a secure 
internet connection.  Data entered into the eCRF must be verifiable against source documents 
at the study site. Any changes to the data entered into the EDC system will be recorded in an automated, secure 
audit trail and is Food and Drug Administration (FDA) Code of Federal Regulations (CFR) Title 21 Part 11 
compliant. 
  
Data entered into the eCRF will be validated as defined in the Data Validation Specifications (DVS). Validation 
includes, but is not limited to, validity checks (for example, missing data, range checks) and consistency checks 
(logical checks between variables) to ensure that study data are accurately reported. Additionally, CRO Data 
Management will perform aggregate data review as defined in the DVS to ensure that the data are complete, 
consistent and reasonable. The electronic edit checks will run continually throughout the course of the study and 
queries reviewed by CRO personnel to assure validity as compared to source records. Manual queries may also 
be entered into EDC by Monitoring or Data Management personnel to address identified discrepancies. 
  
Medical conditions/procedures will be coded using MedDRA and prior and concomitant medications will be 
coded using WHODrug. 
  
At the conclusion of the study, each site will be provided with their subject CRFs in Portable Document Format 
(PDF) for archival.  The CRF PDFs will contain subject data, audit trail information, queries including 
responses, and comments. 
 
 
20 PUBLICATIONS 
All data generated from this study are the property of AOBiome LLC and shall be held in strict confidence 
along with all information furnished by AOBiome.  Independent analyses and/or publication of these data by 
the Investigator or any member of his/her staff is not permitted without prior written consent of AOBiome.   
Any formal presentation or publication of data from this trial will be considered as a joint publication by the 
Investigator(s) and appropriate Sponsor personnel.  Authorship will be determined by mutual agreement. 
Written permission to the Investigator will be contingent on the review by the Sponsor of the methodology and 
statistical analysis and any publication or presentation will provide for nondisclosure of AOBiome confidential 
or proprietary information.  In all cases, the parties agree to submit all manuscripts or abstracts to all other 

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parties at least 30 days prior to submission.  This will enable all parties to protect proprietary information and to 
provide comments based on information that may not yet be available to other parties.   
21 PROTOCOL ADHERENCE 
This protocol defines the study objectives, the study procedures and the data to be collected on study 
participants. Additional assessments required to ensure safety of participants should be administered as deemed 
necessary on a case by case basis. Under no circumstances is an Investigator allowed to collect additional data 
or conduct any additional procedures for any research related purpose involving any investigational drugs under 
the protocol. 
Investigators ascertain they will apply due diligence to avoid protocol deviations. If an Investigator feels a 
protocol deviation would improve the conduct of the study this must be considered a protocol amendment, and 
unless such an amendment is agreed upon by the sponsor and approved by the IRB/IEC and health authorities, 
where required, it cannot be implemented. 
22 CLINICAL TRIAL PROTOCOL AMENDMENTS 
Any protocol amendments will be added as stand-alone documents.  In addition, any and all revisions dictated 
by the amendments will be made in the protocol.  Each time a protocol is amended, a new amended version date 
will be added to the cover page. 
The Investigator should not implement any deviation from, or changes of the Clinical Trial Protocol without 
agreement by the sponsor and prior review and documented approval/favorable opinion from the IRB/EC of an 
amendment. The Investigator is expected to take any immediate action required for the safety of any participant 
included in this study, even if this action represents a deviation from the protocol. In such cases, the reporting 
requirements identified in Section 13 must be followed and the Study Lead. 
23 CONFIDENTIALITY  
All information disclosed or provided by the Sponsor (or any company/institution acting on their behalf), or 
produced during the Clinical Trial, including, but not limited to, the Clinical Trial Protocol, the CRFs, the 
Investigator's Brochure and the results obtained during the course of the Clinical Trial, is confidential. The 
Investigator or any person under his/her authority agrees to undertake to keep confidential and not to disclose 
the information to any third party without the prior written approval of the Sponsor. However, the submission of 
this Clinical Trial Protocol and other necessary documentation to the Ethics Committee (IRB/EC) is expressly 
permitted, the IRB/EC members having the same obligation of confidentiality. The Sub-Investigators shall be 
bound by the same obligation as the Investigator. The Investigator shall inform the Sub-Investigators of the 
confidential nature of the Clinical Trial. The Investigator and the Sub-Investigators shall use the information 
solely for the purposes of the Clinical Trial, to the exclusion of any use for their own or for a third party's 
account. 
 
24 PROPERTY RIGHTS  
All information, documents and Investigational Product provided by the Sponsor or its designee are and remain 
the sole property of the Sponsor. The Investigator shall not mention any information or the Product in any 
application for a patent or for any other intellectual property rights. All the results, data, documents and 
inventions, which arise directly or indirectly from the Clinical Trial in any form, shall be the immediate and 
exclusive property of the Sponsor. The complete verified database will be shared with the Operations 
Committee, which shall have full access to all data. The Sponsor may use or exploit all the results at its own 

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discretion, without any limitation to its property right (territory, field, continuance). The Sponsor shall be under 
no obligation to patent, develop, market or otherwise use the results of the Clinical Trial. As the case may be, 
the Investigator and/or the Sub-Investigators shall provide all assistance required by the Sponsor, at the 
Sponsor's expense, for obtaining and defending any patent, including signature of legal documents.  
 
25 DATA PROTECTION  
The patient's personal data and Investigator's personal data which may be included in the Sponsor database shall 
be treated in compliance with all applicable laws and regulations;  
 
When archiving or processing personal data pertaining to the Investigator and/or to the patients, the Sponsor 
shall take all appropriate measures to safeguard and prevent access to this data by any unauthorized third party.  
26 SPONSOR AUDITS AND INSPECTIONS BY REGULATORY AGENCIES  
For the purpose of ensuring compliance with the Clinical Trial Protocol, Good Clinical Practice and applicable 
regulatory requirements, the Investigator should permit auditing by or on the behalf of the Sponsor and 
inspection by applicable regulatory authorities.  
 
The Investigator agrees to allow the auditors/inspectors to have direct access to his/her study records for review, 
being understood that this personnel is bound by professional secrecy, and as such will not disclose any 
personal identity or personal medical information.  
 
The Investigator will make every effort to help with the performance of the audits and inspections, giving 
access to all necessary facilities, data, and documents. As soon as the Investigator is notified of a future 
inspection by the authorities, he will inform the Sponsor and authorize the Sponsor to participate in this 
inspection.  
 
The confidentiality of the data verified and the protection of the patients should be respected during these 
inspections.  
 
Any result and information arising from the inspections by the regulatory authorities will be immediately 
communicated by the Investigator to the Sponsor.  
 
The Investigator shall take appropriate measures required by the Sponsor to take corrective actions for all 
problems found during the audit or inspections.  
 
27 PREMATURE DISCONTINUATION OF THE STUDY OR PREMATURE CLOSE-OUT OF A 
SITE  
 
27.1 Decided by the Sponsor in the following cases: 
1. In the event the results of the Clinical Trial do not appear to be scientifically convincing to the Sponsor;  
2. If the aim of the Clinical Trial has become outdated or is no longer of interest;  
3. If the information on the product leads to doubt as to the benefit/risk ratio;  

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4. If the Investigator has received from the Sponsor all Investigational Product, means and information 
necessary to perform the Clinical Trial and has not included any patient after a reasonable period of time 
mutually agreed upon;  
5. In the event of breach by the Investigator of a fundamental obligation under this agreement, including 
but not limited to breach of the Clinical Trial Protocol, breach of the applicable laws and regulations or 
breach of the ICH guidelines for GCP;  
6. If the total number of patients are included earlier than expected; In any case the Sponsor will notify the 
Investigator of its decision by written notice.  
 
27.2 Decided by the Investigator  
The Investigator must notify (30 days' prior notice) the Sponsor of his/her decision and give the reason in 
writing. In all cases (decided by the sponsor or by the Investigator), the appropriate Ethics Committee(s) 
(IRB/EC) and Health Authorities should be informed.  
 
28 CLINICAL TRIAL PROTOCOL AMENDMENTS  
 
All appendices attached hereto and referred to herein are made part of this Clinical Trial Protocol. The 
Investigator should not implement any deviation from, or changes of the Clinical Trial Protocol without 
agreement by the sponsor and prior review and documented approval/favorable opinion from the IRB/EC of an 
amendment, except where necessary to eliminate an immediate hazard(s) to clinical trial patients, or when the 
change(s) involves only logistical or administrative aspects of the trial. Any change agreed upon will be 
recorded in writing, the written amendment will be signed by the Investigator and by the Sponsor and the signed 
amendment will be filed with this Clinical Trial Protocol. Any amendment to the Clinical Trial Protocol 
requires written approval/favorable opinion by the IRB/EC prior to its implementation, unless there are 
overriding safety reasons. In some instances, an amendment may require a change to the Informed Consent 
Form. The Investigator must receive an IRB/EC approval/favorable opinion concerning the revised Informed 
Consent Form prior to implementation of the change.  
 
 
 
 
29 REFERENCES 
 
1. Nwankwo T, Yoon SS, Burt V, Gu Q. Hypertension among adults in the US: National Health and 
Nutrition Examination Survey, 2011-2012. NCHS Data Brief, No. 133. Hyattsville, MD: National 
Center for Health Statistics, Centers for Disease Control and Prevention, US Dept of Health and Human 
Services, 2013. 
2. Mozzafarian D, Benjamin EJ, Go AS, et al. Heart Disease and Stroke Statistics-2015 Update: a report 
from the American Heart Association.Circulation. 2015;e29-322. 

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3. Arp DJ, Sayavedra-Soto LA, Hommes NG (2002) Molecular biology and biochemistry of ammonia 
oxidation by Nitrosomonas europaea. Arch Microbiol 178: 250-255. 
4. Chandran K, Stein LY, Klotz MG, van Loosdrecht MCM (2011) Nitrous oxide production by 
lithotrophic ammonia-oxidizing bacteria and implications for engineered nitrogen-removal systems. 
Biochem Soc Trans 39: 1832-1837. 
5. Williams HC, Dellavalle RP, Garner S. Acne vulgaris. The Lancet. 2012;  379(9813): 361-372. 
doi:10.1016/S0140-6736(11)60321-8. 
6. Weller, R., Price, R. J., Ormerod, A. D., Benjamin, N., & Leifert, C. (2001). Antimicrobial effect of 
acidified nitrite on dermatophyte fungi, Candida and bacterial skin pathogens. Journal of Applied 
Microbiology, 90(4), 648-652. 
7. Rico, J., Quiring, J., Hollenbach, S., Enloe, C., & Stasko, N. (2014, August). Phase 2 study of efficacy 
and safety of SB204 in the treatment of acne vulgaris. In Journal Of Investigative Dermatology (Vol. 
134, No. 8, pp. S8-S8). 75 Varick St, 9th Floor, New York, NY 10013-1917 USA: Nature Publishing 
Group. 
8. Park C, Kim G, Patel I, Chang J, Tan X. Improving adherence to acne treatment: the emerging role of 
application software. Clin Cosmet Investig Dermatol. 2014; 7: 65–72. doi: 10.2147/CCID.S46051. 
9. Fitz-Gibbon S, Tomida S, Chiu Bor-Han et. al. Propionibacterium acnes strain populations in the human 
skin microbiome associated with acne. J Invest Dermatol. 2013; 133(9): 2152-2160. doi: 
10.1038/jid.2013.21. 
10. Yentzer BA, Wood AA, Sagransky MJ, et al. An Internet-Based Survey and Improvement of Acne 
Treatment Outcomes. Arch Dermatol. 2011;147(10):1223-1224. doi:10.1001/archdermatol.2011.277. 
11. Weiss J, Shavin J, Davis M. Improving Patient Satisfaction and Acne Severity in Patients With Mild to 
Moderate Acne: The BEST Study. Cutis. 2003; 71(2): 3-4. Available at: 
http://www.ecardiologynews.com/fileadmin/qhi_archive/ArticlePDF/CT/07102S003.pdf. Accessed 
December 4, 2014. 
12. Bergman H, Tsai KY,  Seo SJ, Kvedar JC, Watson AJ. Remote Assessment of Acne: The Use of Acne 
Grading Tools to Evaluate Digital Skin Images. Telemed J E Health. 2009;15(5): 426-30. doi: 
10.1089/tmj.2008.0128. 
13. Burke BM, Cunliffe WJ. The assessment of acne vulgaris - the Leeds technique. Br J Dermatol. 1984; 
111(1): 83-92. doi: 10.1111/j.1365-2133.1984.tb04020.x. 
14. Medansky RS, Handler RM, Medansky DL. Self-evaluation of acne and emotion: A pilot study. 
Psychosomatics. 1981; 22(5) 379-383. doi: 10.1016/S0033-3182(81)73507-2.  
15. Pochi PE, Shalita AR, Strauss JS. Report of the Consensus Conference on Acne Classification. 
Washington, D.C., March 24 and 25, 1990. J Am Acad Dermatol. 1991; 24(3): 495-500. doi: 
http://dx.doi.org/10.1016/S0190-9622(08)80076-X. 
16. Lucky AW, Barber BL, Girman CJ, Williams J, Ratterman J, Waldstreicher J. A multirater validation 
study to assess the reliability of acne lesion counting. J Am Dermatol. 1996; 35(4): 559-569. doi: 
10.1016/S0190-9622(96)90680-5.  
17. Dréno B, Poli F, Pawin H, et al. Development and evaluation of a Global Acne Severity Scale (GEA 
Scale) suitable for France and Europe. J Eur Acad Dermatol Venereol. 2011; 25(1): 43-48. doi: 
10.1111/j.1468-3083.2010.03685.x.  
18. Gupta MA, Johnson AM, Gupta AK. The Development of an Acne Quality of Life Scale: Reliability, 
Validity, and Relation to Subjective Acne Severity in Mild to Moderate Acne Vulgaris. Acta Derm 
Venereol. 1998; 78(6): 451-456. doi: 10.1080/000155598442773 
19. Rizer, RL, Mills, OH, Trookman, NS. (2001) The assessment of acne: a re-evaluation of grading 
strategies. Scientific Poster, Annual Meeting of the Am. Acad. Dermatol. 

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20. Williams, H, Robertson, C, Stewart, A, et al: Worldwide variations in the prevalence of symptoms of 
atopic eczema in the international study of asthma and allergies in childhood. J Allergy Clin 
Immunol 1999 103: 125–138, 
21. Diepgen, TL: Is the prevalence of atopic dermatitis increasing? In: Williams HC ed. Atopic Dermatitis: 
the Epidemiology, Causes and Prevention of Atopic Eczema. 2000 Cambridge: Cambridge University 
Press, 
22. Kobayashi T1, Glatz M2, Horiuchi K3, Kawasaki H4, Akiyama H5, Kaplan DH6, Kong HH2, Amagai 
M4, Nagao K7. Dysbiosis and Staphylococcus aureus Colonization Drives Inflammation in Atopic 
Dermatitis. Immunity. 2015 Apr 21;42(4):756-66. 
23. Roekevisch E1, Spuls PI1, Kuester D2, Limpens J1, Schmitt J3. Efficacy and safety of systemic 
treatments for moderate-to-severe atopic dermatitis: a systematic review. J Allergy Clin Immunol. 2014 
Feb;133(2):429-38. 

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30 APPENDIX A-SCHEDULE OF EVENTS 
 
 
Visit Name  
Screening 
 
Washout 
phase16 
 
Baseline 
Week 2 
Day 14 
Week 4 
Day 28 
Week 6 
Final Visit  
(EOS) 
Day 42 
Early 
Termination 
Visit  
 
Visit Window in days 
-21 to -14 
-14 to -1 
-1 to 0 
+/-1 
+/-1 
+/-1 
 
Informed Consent 
X 
 
 
 
 
 
 
Inclusion/Exclusion 
Criteria21 
Х 
 
X 
 
 
 
 
Demographics 
Х 
 
 
 
 
 
 
Medical History 
Х 
 
 
 
 
 
 
Hanifin and Rajka 
criteria 
X 
 
X 
 
 
 
 
Concomitant 
Medications 
Х 
 
X 
Х 
Х 
X 
X 
Smoking status 
X 
 
 
 
 
 
 
Physical Exam 
X 
 
 
 
 
X 
X 
Body Weight 
X 
 
 
X 
X 
X 
X 
Height 
X 
 
 
 
 
 
 
In office BP and HR13 
X 
 
X 
X 
X 
X 
X 
Urine pregnancy test 
for WOCBP1 
X 
 
 
 
 
 
 
Oral Hygiene 
X 
 
 
X 
X 
X 
X 
EASI 
X 
 
X 
X 
X 
X 
X 
VAS 
X 
 
X 
X 
X 
X 
X 
Skindex 16 
Questionnaire 
X 
 
X 
X 
X 
X 
X 
IGA 
X 
 
X 
X 
X 
X 
X 

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Visit Name  
Screening 
 
Washout 
phase16 
 
Baseline 
Week 2 
Day 14 
Week 4 
Day 28 
Week 6 
Final Visit  
(EOS) 
Day 42 
Early 
Termination 
Visit  
 
Visit Window in days 
-21 to -14 
-14 to -1 
-1 to 0 
+/-1 
+/-1 
+/-1 
 
Clinical chemistry11 
X 
 
 
X 
X 
X 
X18 
Blood for biomarkers12 
 
 
X 
X 
X 
X 
X18 
Skin swabs 
 
 
X 
 
X 
X 
X18 
Microbial 
composition20 
X 
 
X 
 
X 
 
 
IWRS 
 
 
X 
 
 
 
 
Dispense IP to patient 
 
 
X 
X 
 
 
 
Collect Investigational 
Product from patient 
 
 
 
X 
X 
 
 
Investigational Product 
application7 
 
 
X 
Х 
Х 
 
X15 
Investigational Product 
compliance6 
 
 
X 
Х 
Х 
 
X15 
Call to subjects to 
initiate Washout 
phase16 
 
X17 
 
 
 
 
 
Actigraphy watch10 
 
 
X 
 
X19 
 
X15 
Counseling4 
 
X 
X 
Х 
Х 
 
 
Study cleanser 2 
X 
X 
X 
X 
X9 
 
X15 
Moisturizer14 
X 
X 
X 
X 
X9 
 
X15 
 Study diary 3 
 
 
X 
X 
X 
 
X15 
AE monitoring8 
X 
X 
X 
Х 
Х 
X 
X 
T

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1. Urine pregnancy test for WOCBP will be done at Screening 
2. Subjects are to be provided cleanser and moisturizer at Screening visit to start using it at the start of the Washout phase. However, cleanser, 
and moisturizer are available upon request in the event that subjects run out of their initial supplies.  
3. Subjects will be asked to fill out study diary for the duration of the study (Baseline-Day 28) 
4. Subjects will be counseled on the use of study medication, diary and answer any questions subject may have 
5. Skin swabs will be obtained during the office visit and will be kept in the refrigerator until shipped at the end of the study 
6. Weight of an IP kit will be obtained at the at the Baseline visit. Study staff will be asked to weigh 2 bottles without the carton box PRE 
FIRST DOSE. Weight will be recorded in grams. Subjects will be asked to bring both bottles back for the Week 2 and Week 4 visit. Upon 
return for the study visit, 2 bottles will be weighted again.   
7. Subjects will be asked to apply the IP twice daily. First IP application will happen in the office under medical supervision.  
8. AEs will be monitored throughout the trial, starting with the time ICF has been signed.  
9. Cleanser and moisturizer should be returned to the study coordinator. Subjects will be asked to go back to their regular routine.  
10. Actigraphy watch is dispensed at the Baseline visit. Subjects will be asked to wear one watch on each wrist starting at Baseline and through Day 28 visit. 
Subjects are allowed to take watches off during bathing/swimming/showering but will be asked to put watches back once these procedures are completed.  
11. Patients should fast for at least 8 hours before the test. Blood for clinical chemistry will be shipped to the central lab for processing. 
Chemistry, Hematology, HbA1C, Lipid Panel and Angiotensin converting enzyme tests will be done at Screening, Week 2, Week 4 and Week 
6. Serology will only be done at Screening. Kits and lab manuals will be provided by the Cenetron Labs.  
12. Blood samples for biomarkers will be collected and processed on site within 5 minutes of collection. Patients should fast for at least 8 hours 
before the test. Samples will then be frozen onsite and shipped on monthly bases to the Central lab for storage.   
13. Blood pressure readings will be obtained at every visit. Subject should be allowed to rest for > 5 minutes sitting, then serial clinic BP 
measurements and heart (x3) rate will be performed, using the non-dominant arm with a calibrated electronic sphygmomanometer. The 
average of the second and third readings will be documented. 
14. Moisturizer to be used for the duration of the trial will be provided to all subjects starting with the Washout period. Subjects will be asked to 
refrain, if possible, from using any other moisturizer and apply the provided product while they are participating in the trial. Any left over 
moisturizer will be collected at Week 4 visit.  
15. If patient’s Early Termination Visit  happens before Week 4, subject will be asked to return  the Actigraphy watch, cleanser, moisturizer and 
IP to the  study staff. Weigh of an IP will need to be taken.  
16. Subjects will  not need to come in to the clinic in order to start the Washout phase. Once all of the Screening procedures have been completed 
and subject becomes eligible for the study, supplies will be issued to the patient. However, the patient will be asked to hold off  starting with 

CONFIDENTIAL 
 
Amendment 3 
 
Page 61  
the Washout period until results of chemistry and serology become available. At that point, subject will then be notified via the phone that 
they may proceed with the Washout period. Start of the Washout period will be recorded in the CRF by the study staff.  Subjects will be 
instructed to start using the provided cleanser and moisturizer if necessary.  
17. Call to subjects to alert them to initiate the 2-week Washout phase and explain the procedures.  
18. If patient’s Early Termination Visit happens before Week 4, blood for biomarkers, clinical chemistry and skin swab will need to be collected.  
19. Return Actigraphy watches 
20. At Baseline, study staff will examine subject’s body surface area affected by AD and will be asked to identify only one area to be sampled. 
Once the area is identified, study staff will sample this particular treatment area. Sampling area should be noted in subject’s record, so that the 
same treatment area is then sampled at Week 4 visit. 
21. Medical records from patient’s dermatologist or primary care physician to confirm the diagnosis of Atopic Dermatitis are optional. Verbal 
confirmation of the diagnosis present for ³ 12 months is sufficient in order to fulfill this criterion.

CONFIDENTIAL 
 
Amendment 3 
 
Page 62  
 
 
31 APPENDIX B-BODY SURFACE AREA AND IP APPLICATION 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

CONFIDENTIAL 
 
Amendment 3 
 
  
 
32 APPENDIX C EASI QUESTIONNAIRE 
 

EASI guidance December 14
How to Use EASI
The EASI scoring system uses a defined process to grade the severity of the signs of eczema and the
extent affected:
1. Select a body region
Four body regions are considered separately:
2. Assess the extent of eczema in that body region
Each body region has potentially 100% involvement. Using the table below, give each respective
body region a score of between 0 and 6 based on the percentage involvement. Precise
measurements are not required.
% involvement
0
1-9%
10 - 29%
30 - 49%
50 - 69%
70 - 89%
90 - 100%
Region score
0
1
2
3
4
5
6
To aid in your body region grading you can use the diagrams in Appendix 1.
3. Assess the severity of each of
the four signs in that
body region:
Grade the severity of each sign on a scale of 0 to 3:
0
None
1
Mild
2
Moderate
3
Severe
To aid your severity grading, a photographic atlas of suggested categories is available in Appendix 2
.
Remember: Include only inflamed areas in your assessment; do not include xerosis (dryness),
ichthyosis, keratosis pilaris, urticaria, infection (unless there is underlying eczema), or post
inflammatory pigmentation changes.
x
Head and neck
x
Trunk (including the genital area)
x
Upper extremities
x
Lower Extremities (including the buttocks)
9 Take an average of the severity across the
involved region.
9 Half points may be used e.g. 2.5.
9 Palpation may be useful in assessing
edema/papulation as well as lichenification
1.
Erythema
2. Edema/papulation
3.
Excoriation
4.
Lichenification
Further explanations
of these terms can
be found in FAQ’s
(Appendix 4)

EASI guidance December 14
How to record your scores
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EASI score ranges from 0-72.
Body region
Erythema
Edema/
papulation
Excoriation
Lichenification
Area score
Multiplier
Score
Head/neck
(
+
+
+
)
X
X 0.1
Trunk
(
+
+
+
)
X
X 0.3
Upper extremities
(
+
+
+
)
X
X 0.2
Lower extremities
(
+
+
+
)
X
X 0.4
The final EASI score is the sum of the 4 region scores
_______
(0-72)
Two forms of the EASI scoring system are available depending on the age of the patients. The multipliers
for the region score are different in the under 8’s version to reflect the relative proportion of body
regions in young children:
x
Patients 8 years or above
x
Patients under 8 years of age.
The forms can be found in appendix 3.1 and 3.2 and also as word documents on the HOME website
(www.homeforeczema.org)

EASI guidance December 14
Appendix 1: Eczema Area and Severity Index (EASI) - Extent of eczema per body region
Score each region from 0 to 100%
Head & neck
Upper extremities
Trunk
Lower extremities

EASI guidance December 14
Appendix 2: Eczema Area and Severity Index (EASI) –lesion severity atlas

EASI guidance December 14
Appendix 3.1: Eczema Area and Severity Index (EASI) case report form – age <8 years
Area of Involvement: Each body region has potentially 100% involvement. Score 0 to 6 based on the following table:
% involvement
0
1-9%
10 - 29%
30 - 49%
50 - 69%
70 - 89%
90 - 100%
Region score
0
1
2
3
4
5
6
Severity of Signs: Grade the severity of each sign on a scale of 0 to 3:
0
None
1
Mild
2
Moderate
3
Severe
Scoring table:
Body region
Erythema
(0-3)
Edema/
Papulation
(0-3)
Excoriation
(0-3)
Lichenification
(0-3)
Region score
(0-6)
Multiplier
Score per body
region
Head/neck
(
+
+
+
)
X
X 0.2
Trunk
(
+
+
+
)
X
X 0.3
Upper extremities
(
+
+
+
)
X
X 0.2
Lower extremities
(
+
+
+
)
X
X 0.3
The final EASI score is the sum of the 4 region scores:
____________
(0-72)
9 Take an average of the severity across the
involved area.
9 Half points may be used e.g. 2.5.

EASI guidance December 14
v]˘ïXîW
Ìuv^À]˙/v˘~^/}(}urPHô˙
Area of Involvement: Each body area has potentially 100% involvement. Score 0 to 6 based on the following table:
% involvement
0
1-9%
10 - 29%
30 - 49%
50 - 69%
70 - 89%
90 - 100%
Region score
0
1
2
3
4
5
6
Severity of Signs: Grade the severity of each sign on a scale of 0 to 3:
0
None
1
Mild
2
Moderate
3
Severe
Scoring table:
Body region
Erythema
(0-3)
Edema/
Papulation
(0-3)
Excoriation
(0-3)
Lichenification(0-
3)
Region score
(0-6)
Multiplier
Score per body
region
Head/neck
(
+
+
+
)
X
X 0.1
Trunk
(
+
+
+
)
X
X 0.3
Upper extremities
(
+
+
+
)
X
X 0.2
Lower extremities
(
+
+
+
)
X
X 0.4
The final EASI score is the sum of the 4 region scores:
____________
(0-72)
9 Take an average of the severity across the
involved area.
9 Half points may be used e.g. 2.5.

EASI guidance December 14
Appendix 4 - Frequently Asked Questions
What is the difference between edema/papulation and lichenification?
Consider edema/papulation as corresponding to the acute signs of atopic dermatitis that reflect
histological spongiosis. Lichenification are more firm thickened plaques with accentuation of
the skin markings that develop as a result of prolonged scratching or rubbing in chronic disease.
In darker skin types, follicular lichenification may present as firm flat-topped discrete papules.
Grade these chronic lesions as lichenification.
How do I grade prurigo nodules?
Prurigo nodules are larger, deeper lesions as a result of chronic scratching and are graded as
areas of lichenification.
How do I grade erythema in darker skin?
To avoid underestimating inflammation in patients with darker skin tones, take into account the
underlying skin pigment when grading erythema. Often this means increasing your erythema
grade by one level.
Can half-steps be used to assess lesion severity?
The original EASI validation study allowed for half steps. These may be helpful when trying to
average the severity of a parameter over a region. For example, if there are some areas with an
erythema grading of 2 and some areas more consistent with a severity of 3, 2.5 may be a good
choice.
What if most areas in a region are a severity grade of 1, but there are some areas that are a
grade 3?
Attempt to average the severity across the involved areas in that region. If these areas are
close to equal in size, a score of 2 would be most appropriate. If the majority of involved areas
are a grade 1, a score of 1 or 1.5 is more appropriate. Be careful not to score the highest
severity in a region but the average one.
How do I grade xerosis (dryness), ichthyosis and hyperlinear palms?
Unless there is active acute or chronic eczema overlying these findings, they are not included in
the EASI assessment.
How precise should my assessment of eczema extent be?
The region scores, which reflect the extent of eczema, were designed and validated as rough
estimates of the percentage of involved skin. Each region is given a score ranging from 0 to 6,
based on a “ballpark” estimation of extent (see region score table in page 1). If you find it
difficult to provide a rough estimate of disease extent, you can use the schematics in Appendix
1 to guide you. More time-consuming methods for evaluating disease extent such as the rule of
nines or the ‘palm’ method are generally unnecessary, as the EASI was designed to be…easy.

EASI guidance December 14
My patient has responded well to treatment and significantly improved since the last visit.
Should I adjust the grading based on the patient’s relative improvement?
No. The EASI is a static score, meaning that it is done independently at each time point to
reflect current severity. You should grade the EASI per visit regardless of the previous status.
Studies have shown that the EASI score has good responsiveness, meaning that overall it is
sensitive to change and the improvement will be reflected in the total score.
Can the EASI be used in children?
Yes. The EASI is performed in the same method in all age groups, but the calculation of the final
EASI score differs by age. When calculating the EASI, each of the 4 region scores is multiplied by
a constant which reflects the relative contribution of that region to the total body surface area.
For patients 8 years and older the multipliers are 0.1 for the head/neck, 0.2 for the upper
extremities, 0.3 for the trunk and 0.4 for the lower extremities. Below 8 years of age the
head/neck multiplier is increased to 0.2 while the lower extremities multiplier decreases to 0.3,
consistent with the proportions of these regions in childhood. Refer to Appendix 3 for EASI
forms by age.
What happens if a child turns 8 during the course of the study? Which EASI formula should I
use?
There are no clear-cut definitions for this situation. In general, if the study is a short term study
such as an RCT lasting a few months – using the same formula throughout the trial is preferred,
even if the child turns 8 during these months. Keeping the EASI formula consistent in this
scenario can serve to preserve the EASI sensitivity to change (e.g. its change in response to
treatment) without compromising the validity of the score.
In long term studies such as cohort studies lasting a year or longer, it is important to update the
EASI formula based on the physical changes children go through. Switching to the age 8+
formula once a child is older is preferred in that scenario.
What do the terms erythema, edema/papulation, excoriation and lichenification mean?
These are key signs of atopic dermatitis. Recognizing and grading them properly requires
training on the visual and physical exam consistent with these signs. Generally speaking,
erythema is skin redness; edema/papulation refers to an elevation or swelling of the skin (that
should be differed from lichenification below); excoriations are scratch marks that have broken
the skin surface; and lichenification is a leathery thickening of the skin with exaggerated skin
markings.

CONFIDENTIAL 
 
Amendment 3 
 
  
33 APPENDIX D. SKINDEX 16 QUESTIONNAIRE 
 

Skindex16 
ÓMMChren,1997 
Skindex16 - United States/English - Mapi. 
ID6862 / Skindex16_AU2.0_eng-USori.doc 
 
 
 
 
 
 
THESE QUESTIONS CONCERN THE SKIN CONDITION WHICH 
HAS BOTHERED YOU THE MOST DURING THE PAST WEEK 
 
 
 
During the past week, how often 
have you been bothered by: 
 
 
 
 
 
Never 
Bothered 
 
Always 
Bothered 
â 
l 
â  
______________________________ 
 
1. 
Your skin condition itching 
.
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
2. 
Your skin condition burning or stinging 
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
3. 
Your skin condition hurting .
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
4. 
Your skin condition being irritated .
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
5. 
The persistence / reoccurrence of your skin condition .
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
6. 
Worry about your skin condition (For example: that it will 
spread, get worse, scar, be unpredictable, etc) .
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
7. 
The appearance of your skin condition .
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
8. 
Frustration about your skin condition .
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
9. 
Embarrassment about your skin condition .
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
10. 
Being annoyed about your skin condition .
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
11. 
Feeling depressed about your skin condition .
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
12. 
The effects of your skin condition on your interactions 
with others (For example: interactions with family, friends, 
close relationships, etc) .
.
.
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
13. 
The effects of your skin condition on your desire to be 
with people .
.
.
.
.
.
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
14. 
Your skin condition making it hard to show affection 
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
15. 
The effects of your skin condition on your daily 
activities .
.
.
.
.
.
.
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
16. 
Your skin condition making it hard to work or do what 
you enjoy 
.
.
.
.
.
.
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
 
 
Have you answered every item? Yes o No o 

Skindex16 
ÓMMChren, 1997 
Skindex16 - United States/Spanish - Version of 27 Jun 12 - Mapi. 
ID6862 / Skindex16_AU2.0_spa-US.doc 
 
 
 
 
ESTAS PREGUNTAS SE RELACIONAN CON LA AFECCIÓN EN LA PIEL  
QUE MÁS LE HA MOLESTADO DURANTE LOS ÚLTIMOS 7 DÍAS 
 
 
 
Durante los últimos 7 días, ¿con qué 
frecuencia le ha molestado…? 
 
 
 
 
 
Nunca me 
molestó 
 
Siempre 
me molestó 
â 
l 
â  
______________________________ 
 
1. 
La picazón en la afección en la piel .
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
2. 
El ardor o escozor en la afección en la piel 
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
3. 
El dolor en la afección en la piel .
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
4. 
La irritación en la afección en la piel 
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
5. 
La persistencia / recurrencia de la afección en la piel .
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
6. 
La preocupación por la afección en la piel  
(Por ejemplo: de que se extenderá, empeorará, dejará 
cicatriz, sea impredecible, etc.) .
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
7. 
El aspecto de la afección en la piel .
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
8. 
La frustración por la afección en la piel 
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
9. 
La vergüenza por la afección en la piel .
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
10. 
Sentirse fastidiado por la afección en la piel .
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
11. 
Sentirse deprimido por la afección en la piel .
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
12. 
Los efectos de la afección en la piel en sus relaciones 
con los demás (Por ejemplo: relaciones con su familia, 
amigos, relaciones cercanas, etc.) .
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
13. 
Los efectos de la afección en la piel en su deseo de 
estar con otras personas 
.
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
14. 
Que la afección en la piel le dificulte demostrar afecto .
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
15. 
Los efectos de la afección en la piel en sus 
actividades diarias 
.
.
.
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
16. 
Que la afección en la piel le dificulte trabajar o hacer 
lo que disfruta .
.
.
.
.
.
.
.
.
.
.
.
.
   o0     o1     o2      o3     o4      o5     o6 
 
 
 
 
 
¿Ha respondido cada pregunta? 
Sí o No o 

CONFIDENTIAL 
 
Amendment 3 
 
  
34 APPENDIX E. HANIFIN AND RAJKA CRITERIA 
 

Hanifin and Rajka Diagnostic Criteria for Atopic Dermatitis (AD) 
 
 
Major criteria: Must have three or more of: 
1. Pruritus 
 
 
 
 
 
 
 
 
 
 
 
2. Typical morphology and distribution 
 
 
 
 
 
 
 
• 
Flexural lichenification or linearity, erythema, scling, serum-crust 
3. Chronic or chronically-relapsing dermatitis 
 
 
 
 
 
 
4. Personal or family history of atopy (asthma, allergic rhinitis, atopic dermatitis)  
 
 
Minor criteria: Should have three or more of: 
1. Xerosis 
 
 
 
 
 
 
 
 
 
 
2. Ichthyosis, palmar hyperlinearity, or keratosis pilaris 
 
 
 
 
3. Immediate (type 1) skin-test reactivity 
 
 
 
 
 
 
4. Raised serum IgE 
 
 
 
 
 
 
 
 
 
5. Early age of onset 
 
 
 
 
 
 
 
 
 
6. Tendency toward cutaneous infections (especially S aureus and herpes simplex) or 
impaired cell-mediated immunity 
 
 
 
 
 
 
 
7. Tendency toward non-specific hand or foot dermatitis 
 
 
 
 
8. Nipple eczema 
 
 
 
 
 
 
 
 
 
9. Cheilitis 
 
 
 
 
 
 
 
 
 
 
10. Recurrent conjunctivitis 
 
 
 
 
 
 
 
 
11. Dennie-Morgan infraorbital fold 
 
 
 
 
 
 
 
 
12. Keratoconus  
 
 
 
 
 
 
 
 
 
 
13. Anterior subcapsular cataracts 
 
 
 
 
 
 
 
14. Orbital darkening 
 
 
 
 
 
 
 
 
 
15. Facial pallor or facial erythema 
 
 
 
 
 
 
 
16. Pityriasis alba  
 
 
 
 
 
 
 
 
 
17. Anterior neck folds 
 
 
 
 
 
 
 
 
 
18. Itch when sweating 
 
 
 
 
 
 
 
 
 
19. Intolerance to wool and lipid solvents 
 
 
 
 
 
 
20. Perifollicular accentuation 
 
 
 
 
 
 
 
 
21. Food intolerance 
 
 
 
 
 
 
 
 
 
22. Course influenced by environmental or emotional factors 
 
 
 
 
23. White dermographism or delayed blanch 
 
 
 
 
 
 
 

CONFIDENTIAL 
 
Amendment 3 
  
35 APPENDIX F. VISAL ANALOGUE SCALE (VAS) 

CONFIDENTIAL 
 
Amendment 3 
  
36 APPENDIX  G. IGA SCALE 

Investigator Global Assessment Score (IGA) 
 
Score 
Category 
Definition 
0 
Clear 
Minor residual discoloration, no erythema or 
induration/papulation, no oozing/crusting 
1 
Almost Clear 
Trace faint pink erythema with almost no induration/papulation 
and no oozing/crusting 
2 
Mild Disease 
Faint pink erythema with mild induration/papulation and no 
oozing/crusting 
3 
Moderate 
Disease 
Pink-red erythema with moderate induration/papulation and there 
may be some oozing/crusting 
4 
Severe Disease Deep/bright red erythema with severe induration/papulation with 
oozing/crusting