Clinical trial NCT05177328
Conditions: Atopic Dermatitis
Sponsor: National Institute of Allergy and Infectious Diseases (NIAID)
Start date: 2022-07-18; Completion date: 2023-10-13
Enrollment: 21 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
The purpose of this study is to examine the pharmacokinetics or survival of new product containing commensal infection fighting bacteria, on the skin of patients with Atopic Dermatitis (AD), after a single application.

Detailed description:
This study will enroll a minimum of 20 participants, 18-80 years of age, with moderate-to-severe atopic dermatitis (AD) on their ventral arms. A minimum of 13 participants will have a positive Staphylococcus aureus (S. aureus) colonized lesion on both upper extremities. A minimum of 7 participants will have a negative S. aureus colonized lesion on both upper extremities.

The participant's colonization status will be determined from cultures taken during a pre-treatment phase, approximately 7 days prior to receiving study treatment on Day 0. On Day 0, skin swabs will be collected from lesional and non-lesional sites (at least 21cm\^2) on the participant's right and left ventral arms and one non-lesional site on the participant's face. After the skin swab collections, the participant will have ShA9 applied to their right or left ventral arm and placebo applied to their contralateral ventral arm. The assignment of ShA9 and placebo to the dominant and non-dominant arms will be randomized. Additional swabs will be collected 15 minutes, and 1, 2, 4, and 6 hours after the ShA9 and placebo applications on Day 0.

Participants will be asked to return to the clinic 24 hours after receiving their single application and again on Days 3, 10, 17, and 24 for the assessment of adverse events (AEs) and the collection of skin swabs from the identified lesional and non-lesional sites, as needed. After the Day 3 visit, a participant will not be required to complete the Day 10, 17, and 24 visits if their lesional swabs are negative for Coagulase Negative Staphylococcal Species (CoNS).

All randomized participants will complete a final End of Study Phone visit on Day 31 to assess for adverse events and status of their AD.

Arms: arm 1: All participants will receive ShA9 active treatment on one of their ventral arms and placebo on their contralateral arm. The assignment of ShA9 and placebo to the dominant and non-dominant arms will be randomized.
Interventions: intervention 1: S. hominis A 9 Product intervention 2: Placebo
Intervention descriptions: intervention 1: Commensal staph species, phosphate buffered saline, and glycerol. intervention 2: Phosphate buffered saline and glycerol

Protocol documents:
ICF_000.pdf: 
National Institute of Allergy and Infectious Diseases (NIAID) 
Division of Allergy, Immunology and Transplantation (DAIT) 
Informed Consent Template 
 
 
Targeted Investigation of Microbiome Elimination (TIME-1)              Version 2.0 
31Mar2022 
 
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TITLE OF CLINICAL RESEARCH STUDY 
 
ADRN-13: A Pilot Study to Evaluate the Survival of Transplanted Staphylococcus hominis A9 on the Skin of 
Adults with Moderate-to-Severe Atopic Dermatitis 
 
PRINCIPAL INVESTIGATOR 
 
 
Tissa Hata, M.D. 
9500 Gilman Drive #0869 
La Jolla, CA 92093-0869 
Phone: 858-657-8390 
 
YOUR PARTICIPATION IS VOLUNTARY 
 
This study will be explained to you. You will have the opportunity to ask questions. Taking part in this 
study is up to you. If you sign this consent, you agree to take part in the research study. You may change 
your mind at any time. We will give you a copy of this signed consent form to keep. 
 
CONSENT KEY INFORMATION:  
The first pages of this document include a summary of the research study to help you decide 
whether or not to participate.  Detailed information is provided after the key summary 
information. 
 
Why is this research being done? 
 
We would like to learn if a new therapy, a spray containing infection fighting bacteria, can improve 
symptoms of atopic dermatitis (AD) and would like to study how long the infection fighting bacteria will 
survive on the skin after application.  
 
How long will this research last and what will I need to do? 
 
Your participation in this study will last approximately 53 days. 
 
If you agree to participate, you will complete a Screening Visit and the clinical staff will take your vitals 
and record your height and weight. They will ask you questions about your medical history and any 
medications or therapies you’re currently using for your atopic dermatitis. 
 
If you are not currently using any other medications or therapies, skin swab samples will be collected 
from both of your arms, and the clinical staff will give you a moisturizer to use on your arms for about one 
week.  
 
If you are using other medications or therapies, you will be asked if you are willing to stop using the 
medications or therapies to continue in the study. Depending on the medication or therapy, you will need 
to stop use 7 to 28 days prior to the next study visit.  If you agree to stop, you will be asked to return to 
the clinic after you are no longer using the medications or therapies, to have your skin swabs collected.  
 
A few days later, you will be contacted regarding your swab results and eligibility to continue in the study. 
If eligible, you will be asked to return to the clinic for a Treatment Visit about 1 week after your skin swabs 
were collected.  During the visit, skin swabs will be collected from your arms and face prior to receiving 
treatment. A member of the clinical staff will then apply the therapy spray to either your right or left arm 


National Institute of Allergy and Infectious Diseases (NIAID) 
Division of Allergy, Immunology and Transplantation (DAIT) 
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Targeted Investigation of Microbiome Elimination (TIME-1)              Version 2.0 
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and a placebo spray (spray without the bacteria) to your other arm. You and your doctor will not know 
which spray is applied to each arm. You will be asked to remain in the clinic for about 6 hours, to have 
additional skin swabs collected. 
 
You will be asked to return to the clinic for up to 5 additional visits (24 Hours, Day 3, Day 10, Day 17, Day 
24) following your Treatment Visit. During each visit, skin swabs will be collected from your face and arms. 
You will return to clinic for 24 Hour and Day 3 Visits. Based on the amount of bacteria still present on your 
skin after the Day 3 Visit and each subsequent visit, you may be told you do not have to return for the 
Day 10, Day 17, and/or Day 24 Visits. All participants will be asked to complete a Day 31 Final Phone Visit.  
 
Is there any way being in this research can hurt me? 
 
All of the study procedures that will be conducted on you have little to no risk, with one exception. The 
application of the new therapy has a small risk of skin infection. The likelihood of participants developing 
a skin infection is low.  
 
The risks for you from the study procedures are discussed in detail in Section 3, “Risks and/or 
Discomforts.” 
 
Will being in this research study help me in any way? 
 
There may or may not be a direct benefit to you from being in the study.  
 
What other choices do I have besides taking part in this research? 
 
You do not have to take part in this research study. You can still receive treatment for your atopic 
dermatitis at <Insert Site Name>. 
 
 
DETAILED CONSENT INFORMATION 
The rest of the consent document includes detailed information about this study.   
 
1. INTRODUCTION/BACKGROUND 
 
Atopic dermatitis, also called eczema, is a disease with dry, scaly, itchy skin. People with atopic dermatitis 
often experience complications of recurrent bacterial and viral skin infections. They experience recurring 
infections with a specific bacterium, Staphylococcus aureus, or Staph infections. The viruses, bacteria, and 
fungi, which cause the infections, interact with the immune system and trigger worsening of atopic 
dermatitis. This defect in the immune system’s response creates a cycle of skin infection and worsening of 
atopic dermatitis. Standard treatment of bacterial infections includes antibiotics removing good and bad 
bacteria from the skin. Prolonged use of antibiotics also generates resistant bacteria, making the 
treatment ineffective.  
 
2. STUDY COMPONENTS 
 
This study is funded by the National Institute of Allergy and Infectious Diseases (NIAID). 
 
This study will enroll approximately 20 participants, 18 to 80 years of age, from one center in the United 
States. All participants will have atopic dermatitis.  
 


National Institute of Allergy and Infectious Diseases (NIAID) 
Division of Allergy, Immunology and Transplantation (DAIT) 
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If you agree to participate in this study, you will be required to comply with UCSD’s current institutional 
policies and procedures for COVID-19. These requirements will be communicated to you at the time of 
visit scheduling and prior to any scheduled appointments. 
 
If you choose to participate in this study, you will have one dose of therapy with infection fighting bacteria 
applied to one of your arms and a placebo (product without the bacteria) applied to your other arm. You 
will not be able to choose which product is applied to each arm. Neither you nor your study doctor will 
know which product was applied to each arm. You will be randomly assigned, much like flipping a coin, to 
receive either the spray with bacteria or the placebo on your dominant arm.  
 
Your participation in the study will last approximately 53 days and require 8-9 visits to <Insert Site Name>. 
If you are using medications/therapies at your first clinic visit, you may be asked to return for an 
additional clinic visit after you have stopped using the medications/therapies. The study doctor will review 
medications/therapies you are currently using to decide if they are safe to stop using while participating 
in the study. You must agree to stop using any prohibited medications/therapies while participating in the 
study.    
 
If you are a female participant of reproductive potential and sexually active, you will be asked to use 
adequate contraception, over the course of your participation in the study. Acceptable methods of 
contraception for female participants include total abstinence, oral contraceptives, intrauterine device 
(IUD), barrier method with spermicide, surgically sterilized partner, Depo-Provera, Norplant, NuvaRing, or 
hormonal implants.  
 
If you agree to participate in this study, you will be given Cetaphil lotion to use for about one week on 
your arms prior to confirming your eligibility to continue and receive the therapy product. After this, we 
would like you to use the Cetaphil lotion we will provide on your arms where we apply the treatment and 
on the area we swab on your face as needed for dry skin, for the duration of the study.  We will also give 
you Dove moisturizing soap for use when showering over the course of your participation. You will not be 
permitted to use the Dove soap on your arms, and you will be asked to refrain from swimming in 
chlorinated pools/hot tubs through your last clinic visit. We will give you a handcard that summarizes all 
of these reminders.  
 
The following procedures will occur during this study: 
 
Screening Visit (Day -14 to Day -7) 
Your initial visit to the center will take approximately 1.5 hours to complete. During the visit, the following 
will occur: 
• 
Consent:  We will explain the study to you. You will be given time to read this consent form and 
ask questions. Once all of your questions are answered, you will sign this consent form if you 
want to participate in the study.  
We may also talk to you over the phone about this study or send this informed consent 
document to you before you come into the clinic for an in-person visit. If you receive it before 
the clinic visit, please review the entire document. Do not sign and date the document until after 
we contact you to discuss the document. We may be able to discuss the consent by video or 
phone. If you agree to join the study, you can sign and date the document during our discussion. 
If you provide a signature by signing and dating with an actual pen, you will need to mail it back 
to us or bring it during your visit to the clinic. If you provide an electronic signature, we will give 
you instructions on how to return the form electronically. 
• 
Questionnaires: You will answer some questions about your contact information and some 
questions to determine if you are eligible for this study. These questionnaires are not tests. There 


National Institute of Allergy and Infectious Diseases (NIAID) 
Division of Allergy, Immunology and Transplantation (DAIT) 
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are no right or wrong answers to the questions. If you do not understand certain questions, you 
may ask the study staff members for more information.  
• 
Physical Exam: You will receive a physical exam by a study doctor or another qualified staff 
member. The purpose of the exam is to make sure that you meet the health requirements for 
the study. During the exam, a member of the study staff will ask you questions about your health 
and atopic dermatitis.  
• 
Medication/Therapy Review: A member of the study staff will ask about the medications you 
have recently taken and any therapies you have used. The use of certain medications may mean 
that you will need to return for an additional visit to continue your participation. 
• 
Pregnancy Status and Test: If you are a female participant and do not self-report as pregnant, 
you will be asked to give a small sample of urine for a pregnancy test. Results of the pregnancy 
test will be reported to you. If your pregnancy test is positive, you will not be allowed to 
participate in the study.   
• 
Vital Signs, Height, and Weight: A member of the study staff will check your vital signs, 
(temperature, heart rate, respiration, and blood pressure), height, and weight.  
 
At the end of the Screening Visit, if you meet all other eligibility requirements and have not taken or 
received certain medications and therapies for your atopic dermatitis, you may be able to complete your 
Pre-treatment Visit on the same day as the Screening Visit.  
 
If you are currently using other medications or therapies, wish to continue participating in the study, and 
meet all other eligibility requirements, the study team will schedule your next study visit after you have 
stopped using the medications/therapies. The study doctor will review your specific 
medications/therapies and confirm the length of time you have to stop use before scheduling your next 
visit. This length of time could be up to 28 days or as short as 7 days.  
 
If you do not meet all eligibility requirements or you are unable or unwilling to stop the use of certain 
medications/therapies, you will not be allowed to participate in the study.  
 
Pre-treatment Visit (Day -7) 
After the Screening Visit, if you are eligible, you will be asked to complete the Pre-treatment Visit. Based 
on your medication/therapy use, this visit may be on the same day as your Screening Visit or 1-2 weeks 
later. This visit will take about 1 hour if conducted on a different day than Screening and about 30 minutes 
if conducted on the same day. During the visit, the following will occur: 
• 
Questionnaires 
• 
Physical Exam 
• 
Medication/Therapy Review 
• 
Vital Signs 
• 
Pregnancy Status (Female participants only) 
• 
Skin Swab Collection: A member of the study staff will take pictures of your arms. A total of 8 
swab samples will be collected from areas of affected and unaffected skin. The samples will be 
collected by rolling a sterile swab over the skin. Collection of each swab will take about one 
minute and is painless. Your skin swabs and pictures will be labeled with a number only, and not 
your name. The swabs will be used to look at bacteria growing on your skin. 
• 
Moisturizer Use: The study team will give you a moisturizer to use at least twice a day on your 
arms for a total of about 7 days before your next visit.  
 
A few days following your Pre-treatment Visit, a member of the study team will contact you about your 
swab results and eligibility to continue with study treatment. If your results meet eligibility for a group 
that has already enrolled enough participants, you may not be able to continue in the study.  


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Treatment Visit (Day 0) 
Seven days after your Pre-treatment Visit, if you are eligible you will be asked to return to the clinic to 
have the therapy applied to one of your arms and placebo applied to the other. This visit will take 
approximately 7 hours. During the visit, the following will occur: 
• 
Questionnaires 
• 
Physical exam 
• 
Medication/Therapy Review 
• 
Pregnancy Test (Female participants only) and pregnancy status for partners of participants, as 
applicable 
• 
Vital Signs 
• 
Skin Swab Collection: 4 swabs will be collected from each arm, and 2 swabs will be collected 
from your face (10 swabs total) 
• 
Therapy Application: A study staff member, while wearing gloves, will apply therapy spray to one 
of your arms and the placebo spray to your other arm.  
• 
Skin Swab Collection: After therapy and placebo are applied to your arms, 8 swabs (4 from each 
arm) will be taken at each of the following time points:  15 minutes, 1, 2, 4, and 6 hours. 
 
Follow-Up Visits (24 Hour and Days 3, 10, 17, and 24) 
After the Treatment Visit, you will be asked to return to the clinic for up to 5 follow-up visits at 24 hours, 
and 3, 10, 17, and 24 days post treatment. Each visit will take approximately 45 minutes. During each visit, 
the following will occur: 
• 
Physical Exam 
• 
Medication/Therapy Review 
• 
Pregnancy Status (Female participants and partners of participants, as applicable) 
• 
Vital Signs 
• 
Skin Swab Collection: 4 swabs from each arm and 2 swabs from your face will be collected at the 
24 Hour Visit and Day 3 – 24 Visits  
 
After the Day 3 Follow-Up Visit and each subsequent visit, the study staff will call you prior to your next 
scheduled visit if you do not need to return for the remaining follow-up visits (Day 10, Day 17, and/or Day 
24).  
 
End of Study Phone Visit (Day 31) 
A study staff member will call you approximately one month after your 24 Hour Follow-Up Visit. The call 
will take approximately 15 minutes. You will be asked about your atopic dermatitis, any new symptoms 
you’re experiencing, any medications/therapies you have used since your last visit, and whether or not 
you (female participants) or your partner have tested positive to a pregnancy test since your last visit to 
the center. 
 
Unscheduled Visit 
In the event you experience increased disease activity, signs and symptoms of an infection (pain or 
swelling at the site of treatment, fever, chills, or night sweats), or other concerns arise between your 
study visits, you may contact the clinical center. An emergency handcard summarizing these symptoms 
and the clinic phone numbers will be provided to you. Study staff will assess if you need to return to clinic 
for an additional visit. During the visit the following will occur: 
• 
Physical Exam 
• 
Medication/Therapy Review 
• 
Pregnancy Status (Female participants and partners of participants, as applicable)  
• 
Vital Signs 
• 
Skin Swab Collection: total swabs collected will be at the discretion of the doctor  


National Institute of Allergy and Infectious Diseases (NIAID) 
Division of Allergy, Immunology and Transplantation (DAIT) 
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• 
Blood Collection: Approximately 0.5 teaspoon of blood may be collected from your arm to check 
your blood count. Additional blood, approximately 2 teaspoons per culture, may be collected at 
the discretion of the doctor. All of the blood can usually be collected with 1 needle stick. Your 
blood sample will be labeled with a number only, and not your name.   
 
Should you experience increased disease activity or signs and symptoms of an infection and seek care at 
an outside facility, you should take the emergency handcard with you and provide it to the physician. The 
card will explain that you are in a clinical trial and include contact information for <Insert PI Name> and 
the 24 Hour On-Call physician at <Insert Site Name>. The card will also provide helpful information to the 
physician so he/she can treat your symptoms.  
  
3. RISKS and/or DISCOMFORTS 
 
Treatment and procedures in this research study may involve risks that are not possible to predict. You 
will be informed of any new risks that may be identified during the study. Below is a description of the 
risks we know about for each procedure. Please ask your study doctor or the research staff to explain any 
procedures or risks that you do not understand. 
 
• 
Questionnaires: You may find that some of the questions are too personal. You may refuse to 
answer any questions that make you feel uncomfortable. There is also a possibility that your 
answers may be read by others outside of the study. Your name is not put on the questionnaires.  
• 
Physical Exam: There are no known risks for the physical exam. 
• 
Medication/Therapy Washout: Stopping medications and/or therapies prior to sample collection 
and during your study participation may cause worsening of your atopic dermatitis.  
• 
Pregnancy Test: There are no known risks for the pregnancy test. 
• 
Vital Signs, Height, and Weight: There are no known risks for having your vital signs, height, and 
weight checked. 
• 
Skin Swab Collection: There are no known risks for the skin swab collection.  
• 
Therapy Spray Application: Although the likelihood of participants developing a skin infection is 
low, a skin infection may occur where bacteria are applied to the skin. In order to reduce this 
risk, product will not be applied to your skin if you have evidence of skin cracks or breaks. 
Product will not be applied to your hands, and the study staff will wear gloves when applying the 
product, to reduce the chances of spreading bacteria onto other surfaces or people. If product is 
applied to other areas of the body, including eyes, nose, mouth, or genital area, there may be a 
higher risk of irritation or infection.  
• 
Blood Collection: The risks of having blood taken may include pain, bleeding, or bruising. Some 
people may experience lightheadedness, nausea, or fainting. There is a potential for slight 
psychological stress from the procedure. If psychological stress is too much in the opinion of the 
participant or study staff, the procedures will be halted. 
• 
There is a risk of loss of confidentiality of your information. Additional details regarding your 
confidentiality are included in Section 14. Confidentiality.  
 
4. POTENTIAL BENEFITS    
 
If you agree to take part in this study, there may be no direct medical benefit to you. 
Information learned from this study may someday benefit the future treatment and care of people with 
atopic dermatitis.  
 


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Division of Allergy, Immunology and Transplantation (DAIT) 
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5. ALTERNATIVES TO PARTICIPATION 
 
There are other options available for you if you decide not to enroll in this study. You may receive 
treatment from a doctor without being in this study. You may choose to enroll in another study. You do 
not have to participate in this study in order to receive treatment at <Insert Site Name>. 
 
6. NEW FINDINGS  
 
The study doctor will tell you about any new information that may affect your willingness to continue in 
this study. 
 
7.  VOLUNTARY WITHDRAWAL FROM STUDY  
You may decide not to take part or to leave the study at any time. If you decide not to participate or to 
leave the study, there is no penalty or loss of benefits in your routine medical care or any other benefit(s) 
that you would otherwise be entitled to receive.   
 
If you leave the study prior to completion, we may ask you to complete a final assessment by phone, to 
confirm that any ongoing signs and symptoms related to your participation in the study are resolved and 
to answer any questions you may have about your atopic dermatitis.  
 
If you decide to leave the study or if you decide you do not want us to keep your contact information, you 
can call <Insert Site Contact Number> to remove your information from our list. You can also write us at 
<Insert Site Address> to let us know that you want to leave the study. 
 
8. REASONS WHY YOU MAY BE TAKEN OFF STUDY WITHOUT YOUR CONSENT 
 
You may be removed from the study without your consent at any time.  Reasons why you may be 
removed from the study include, but are not limited to, the following:      
• 
The study doctor feels it is not in your best interest to continue in this study.   
• 
You are unable to complete required study procedures, due to the development of a new 
disease, becoming pregnant, or starting treatment(s) not allowed for study participants.  
• 
The study is stopped by <Insert Site Name>, the National Institute of Allergy and Infectious 
Diseases, or other health authorities. 
• 
You do not comply with study procedures or follow study rules.  
 
9.  PREGNANCIES, BREASTFEEDING AND BIRTH CONTROL  
 
You cannot participate in this study if:   
• 
You are currently pregnant or breastfeeding. 
• 
You plan to get pregnant in the next 2 months. 
 
Treatments and procedures involved in this research project may involve unexpected risks to your unborn 
or nursing child.  If you are female and of childbearing potential, a pregnancy test will be performed prior 
to your enrollment in this study.  
 
If you are a female of child-bearing potential, you must agree to use birth control for the duration of your 
participation in the study.  The acceptable methods of birth control are as outlined above in Section 2, 
Study Components.  
 


National Institute of Allergy and Infectious Diseases (NIAID) 
Division of Allergy, Immunology and Transplantation (DAIT) 
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31Mar2022 
 
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If you are a female and become pregnant while participating in this study, or if you suspect that you have 
become pregnant, please contact < Insert PI Name> immediately.   
 
10. COSTS TO THE SUBJECT (YOU) 
 
There will be no charge to you or your health insurance company for any costs for the study therapy or 
placebo. There will be no charge to you or your health insurance company for any costs which are directly 
related to this study’s procedures:  
• 
Skin Swab Collection  
• 
Blood draw 
The normal costs related to your atopic dermatitis care during the course of the study will be up to you 
and/or your insurer: 
• 
Physician visits outside of study visits 
• 
Emergency room or urgent care visits 
• 
Hospitalizations 
 
11. PAYMENTS (REIMBURSEMENT) 
 
You will be compensated for your time and effort to participate in this study. 
 
At the Screening Visit, you will receive $40 for completing the activities, which include the questionnaires, 
pregnancy testing, and physical exam. If you complete the Pre-treatment Visit, you will receive an 
additional $50.  
 
You will receive $150 for completing the Treatment Visit. 
 
If you complete the 24 Hour, Day 3, Day 10, Day 17, and Day 24 Follow-Up visits, you will receive $50 per 
visit. 
 
You will receive $10 for completing the End of Study Phone Visit. 
 
In the event you are asked to return to clinic for an Unscheduled Visit, due to experiencing increased 
disease activity, showing signs and symptoms of an infection (pain, swelling, redness or tenderness at the 
site of treatment, fever, chills, or night sweats), or other concerns, you will receive $50 for each visit.  
 
If you choose to or are asked to leave the study before it is finished, you will only be compensated for the 
activities you complete. 
 
It is important to know that payment for participation in this study may be taxable income.  
 
12. RESEARCH-RELATED INJURY 
 
If you are injured or become ill because of taking part in this study, it is important to tell your study doctor 
< Insert PI Name>.   Emergency medical treatment will be available to you, through <Insert Site Name>. 
You or your health insurance provider will be billed for the payment for any treatment you require as a 
result of a study-related injury. No other form of reimbursement is available. <Insert Site Name> and the 
National Institutes of Health, including the Division of Allergy, Immunology, and Transplantation do not 
have programs to pay if you are hurt or have other bad results that are not the fault of the study doctors. 
You may contact <Insert name of Research Staff Office> at <Insert Office Contact Number> for more 
information or to report study-related injuries.  


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In case of injury resulting from this study, you will not lose any legal rights by signing 
this form. 
 
13. DISCLOSURE OF FINANCIAL CONFLICT OF INTEREST  
 
Protocol Chair, Dr. Richard Gallo of the University of California – San Diego, is a co-founder of MatriSys 
Bioscience, Inc. (MatriSys) and has previously served as Chair of the company’s Scientific Advisory Board. 
Dr. Gallo holds equity within the company and as Protocol Chair, Dr. Gallo helped design this study. 
MatriSys is working towards developing therapeutic products containing live bacteria to treat atopic 
dermatitis. MatriSys is not involved in the manufacture of the product to be used in this clinical study. 
 
14. CONFIDENTIALITY  
 
Your medical and research records will be confidential to the extent permitted by law.  Efforts will be 
made to keep your personal information private.  However, we cannot guarantee complete 
confidentiality.   
 
You will be identified by a code, and personal information from your records will not be released without 
your written permission.  You will not be identified in any publication or in the sharing of your data about 
this study.  After the study is completed, the data may be placed in a central storage location.  These data 
will not include your name or other information that can identify you.  The purpose is to make study data 
available to other researchers.  Information from this study may be put into databases along with 
information from other studies.  There are different kinds of databases; some are publicly accessible, and 
some are restricted.  Anyone on the internet can access publicly accessible databases.  Only researchers 
who apply and are approved can access restricted databases.  Traditionally used identifying information 
about you (such as name, phone number, address) will NOT be included or shared with others. 
 
As an NIH funded study, you are further protected through a policy that prevents the study doctor from 
releasing any sensitive information about you that may identify you.  This does not prevent you or a 
family member from voluntarily releasing information about this research. 
 
Organizations that may look at and/or. copy your medical records for research, quality assurance, and 
data analysis include:   
1. The National Institute of Allergy and Infectious Diseases (NIAID) sponsor of the research  
2. NIAID representatives, agents, employees, contractors, and other persons assisting in 
conducting, monitoring or analyzing the study.  This includes the study monitors from Rho, Inc., 
<Insert local IRB >, Principal Investigator, and research staff at <Insert Site Name> 
3. The U.S. Food and Drug Administration  
4. Other State and Local health authorities   
A description of this clinical trial will be available on http://www.ClinicalTrials.gov, as required by U.S. law.  
This web site will not include information that can identify you.  At most, the Web site will include a 
summary of the results.  You can search this Web site at any time. 
 


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15. PROBLEMS OR QUESTIONS 
 
 I can call …          
 At …         
 If I have questions or concerns about 
During <Insert Regular Clinic Hours> 
<Insert PI Name>  
Principal Investigator   
Phone:      <Insert Phone>   
Pager:       <Insert Pager>   
Beeper # 
General questions about the study 
Research- related injuries or emergencies 
Any research-related concerns or complaints  
Symptoms in between my regular scheduled study visits 
Outside of Regular Clinic Hours 
On Call Physician 
Phone:      <Insert Phone>   
Pager:       <Insert Pager>   
Beeper # 
Research- related injuries or emergencies 
Any research-related concerns or complaints  
Symptoms in between my regular scheduled study visits 
<Insert Research Staff Office Number>   Phone:      <Insert Phone>   
Pager:       <Insert Pager>   
Beeper # 
General questions about the study 
Research- related injuries or emergencies 
Any research-related concerns or complaints 
<Insert IRB> 
Phone:       <Insert Phone>     
Rights of a research subject 
Any research-related concerns or complaints 
If investigator/study contact cannot be reached 
If I want to speak with someone other than the 
Investigator, study contact, or research staff 
16. FUTURE USE OF YOUR DATA/BIOLOGIC MATERIALS 
 
Information about you, including your biospecimens, collected for this study may be shared with other 
researchers.  It may also be used for other research studies.  We will make sure that your identity cannot 
be linked to the information we share; your data and biospecimens will be coded so that other 
investigators will not have access to your personal information.  We will not ask you for additional 
permission before sharing the information. 
 
We are asking your permission to store unused skin swab samples and secondary samples created from 
your skin swabs collected during the course of this study to be used in the future for tests that aren’t yet 
planned.  No human DNA analysis will be performed on these samples.    
 
The results of tests performed on stored samples or reports resulting from the analysis of your samples will 
not be given to you or your doctor, and they will not be put in your medical record.  They will not identify 
you and will not affect your routine medical care. 
 
There is no benefit to you from the storage of samples and information.  The purpose of storage and 
sharing data is to make information available for use in health research and to share what is stored with 
other researchers.  Collecting, storing, and sharing information and making it available for other studies 
may help people in the future.   Coded information put into databases together with other stored 
information from many studies conducted in different places allow researchers to study the combined 
information and learn even more about health and many different diseases.    
 
Samples will be stored at the University of California – San Diego.  If you decide to allow storage, your 
samples and information may be stored for an unknown length of time. 
 
Although your stored research samples will not be sold, the information obtained from the research 
performed on your samples may in the future lead to the development of commercial products.  You will 
not receive any money from research using your stored samples and information. 
There may be unknown risks associated with the storage and analysis of samples or the information 
resulting from the analysis of your samples.   


National Institute of Allergy and Infectious Diseases (NIAID) 
Division of Allergy, Immunology and Transplantation (DAIT) 
Informed Consent Template 
 
 
Targeted Investigation of Microbiome Elimination (TIME-1)              Version 2.0 
31Mar2022 
 
Page 11 of 12 
You can change your mind at any time and ask to have your samples destroyed.  This request should be 
made in writing to the study doctor.  If you make this request, all remaining stored samples will be 
destroyed.  However, the results of any previous tests using your stored samples will be used.    Your 
decision regarding the storage of samples or the information resulting from the analysis of your samples 
will not affect your ability to participate in this study.   
 
I agree to the storage and sharing of skin swabs or any secondary samples created from skin swabs and 
information resulting from the analysis of my samples for other tests not currently planned.  
 
 
   Yes       
  No    
 
 
 
                                    _______________________ 
 
 
      Initials of Research Subject 
 
 
 


National Institute of Allergy and Infectious Diseases (NIAID) 
Division of Allergy, Immunology and Transplantation (DAIT) 
Informed Consent Template 
 
 
Targeted Investigation of Microbiome Elimination (TIME-1)              Version 2.0 
31Mar2022 
 
Page 12 of 12 
17. PARTICIPANT SIGNATURE PAGE 
 
Please sign below if you agree to take part in this study.  
  
• 
You have read the informed consent and had it explained to you 
• 
You were given the opportunity to ask questions about the information 
• 
You understand the risks and benefits of this study as explained in this consent form 
• 
You voluntarily agree to take part in the study 
 
 
 
_________________ 
         
_____________________      
 
______________ 
Research Participant’s Name 
Research Participant’s Signature                           Date 
(Typed or printed) 
 
 
 
Signature of person explaining and obtaining the consent: 
 
 
 
____________________ ____________________  
_________________ 
Name and Title  
Signature 
 
 
 
Date 
(Typed or printed)   
 
  
(NOTE:  This consent form with the original signatures MUST be retained on file by the principal 
investigator.  A copy must be given to the research participant.  A copy should be placed in the research 
participant’s medical record, if applicable.)      
 
 
 
 
 
 
 
 
 
 
 
 
 

SAP_002.pdf: 
DAIT/Rho STATISTICAL ANALYSIS PLAN For INTERIM 
and FINAL ANALYSIS  
12 March 2024 
 
 
A Pilot Study To Evaluate The Survival Of Transplanted Staphylococcus 
Hominis A9 On The Skin Of Adults With Moderate-To-Severe Atopic 
Dermatitis  
VERSION: 
2.0 
DATE: 
12 March 2024 
SPONSOR: 
 
National Institute of Allergy and Infectious Diseases – NIH 
5601 Fishers Lane 
Rockville, MD 20852 
Telephone: (301) 496-5717 
Fax: (301) 402-3573 
 
 
PREPARED BY: 
Rho 
Rho Building 
 
2635 E NC Hwy 54 
Durham, NC 27713 
Telephone: (919) 408-8000 
Fax: (919) 408-0999 
 
 
 
 


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DAIT/Rho STATISTICAL ANALYSIS PLAN For INTERIM 
and FINAL ANALYSIS 
ACKNOWLEDGMENT AND SIGNATURE SHEET 
 
A Pilot Study To Evaluate The Survival Of Transplanted Staphylococcus 
Hominis A9 On The Skin Of Adults With Moderate-To-Severe Atopic 
Dermatitis 
Approved: 
__________________________________ 
Date: 
_______________ 
 
Richard Gallo, MD, PhD 
 
 
 
Protocol Chair 
University of California San Diego 
 
 
Approved: 
__________________________________ 
Date: 
_______________ 
 
Tissa Hata, MD 
 
 
 
Protocol Co-Chair 
University of California San Diego 
 
 
Approved: 
__________________________________ 
Date: 
_______________ 
 
 
 
 
 
NIAID Medical Monitor 
 
 
 
NIH/NIAID 
 
 
Approved: 
__________________________________ 
Date: 
_______________ 
 
 
 
 
 
Principal Research Scientist 
Rho, Inc. 
 
 
Approved: 
__________________________________ 
Date: 
_______________ 
 
 
Biostatistician 
Rho, Inc. 
 
 
 


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Version: 2.0 
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Document History 
 
Version 
Date 
Change(s) 
1.0 
14 AUGUST 2023 
Initial Document 
2.0 
12 MARCH 2024 
Updated the planned analysis for secondary endpoint #1.  
Updated and clarified which analysis populations will be used for the 
planned analyses. 


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Table of Contents 
 
1. PROTOCOL SYNOPSIS (VERSION 3.0) ......................................................................... 6 
2. INTRODUCTION ............................................................................................................ 11 
3. GENERAL ANALYSIS AND REPORTING CONVENTIONS .......................................... 12 
4. ANALYSIS SAMPLES ................................................................................................... 13 
4.1 Safety Sample ..................................................................................................... 13 
4.2 Modified Intent-to-Treat (mITT) Sample ............................................................... 13 
4.3 Per-Protocol (PP) Sample.................................................................................... 13 
4.4 Interim Analysis Sample ...................................................................................... 13 
5. STUDY PARTICIPANTS ................................................................................................ 14 
5.1 Disposition of Participants .................................................................................... 14 
5.2 Demographic and Other Baseline Characteristics ................................................ 14 
5.3 Medical History .................................................................................................... 14 
6. STUDY OPERATIONS ................................................................................................... 14 
6.1 Protocol Deviations .............................................................................................. 14 
6.2 Treatment Adherence .......................................................................................... 14 
7. ENDPOINT EVALUATION ............................................................................................. 15 
7.1 Overview of Efficacy Analysis Methods ................................................................ 15 
7.1.1 Handling of Dropouts or Missing Data .......................................................... 15 
7.1.2 Multicenter Studies ....................................................................................... 15 
7.1.3 Assessment Time Windows ......................................................................... 15 
7.2 Reporting of Swab Results .................................................................................. 15 
7.3 Primary Objective ................................................................................................ 16 
7.3.1 Computation of the Primary Endpoint ........................................................... 16 
7.3.2 Primary Analysis of the Primary Objective .................................................... 16 
7.3.3 Supportive Analyses of the Primary Endpoint ............................................... 16 
7.4 Secondary Objectives .......................................................................................... 17 
7.4.1 Secondary Endpoints ................................................................................... 17 
7.4.2 Analyses of Secondary Endpoints ................................................................ 18 
7.5 Exploratory Objectives ......................................................................................... 18 
7.5.1 Exploratory Endpoints .................................................................................. 18 
7.5.2 Analyses of the Exploratory Endpoints ......................................................... 19 
8. SAFETY EVALUATION ................................................................................................. 19 
8.1 Overview of Safety Analysis Methods .................................................................. 19 
8.2 Adverse Events ................................................................................................... 20 
8.3 Deaths and Serious Adverse Events ................................................................... 20 
8.4 Vital Signs, Physical Findings, and Other Observations Related to Safety ........... 20 
8.4.1 Vital Signs .................................................................................................... 20 
8.4.2 Pregnancy Assessment ............................................................................... 20 
9. OTHER ANALYSES ....................................................................................................... 21 
9.1 Use of Medications .............................................................................................. 21 
10. INTERIM ANALYSES AND DATA MONITORING ....................................................... 21 
10.1 Interim Safety Reporting to the Data and Safety Monitoring Board .................... 21 
10.2 Interim Analysis of Primary Endpoint ................................................................. 21 
11. SAMPLE SIZE CONSIDERATIONS ............................................................................. 22 
12. CHANGES TO THE ANALYSES PLANNED IN THE PROTOCOL .............................. 22 
13. REFERENCES ............................................................................................................. 23 
14. APPENDICES .............................................................................................................. 24 
14.1 Study Flow Chart ............................................................................................... 24 
14.2 Schedule of Events ............................................................................................ 25 
 
 


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List of Tables 
 
Table 1 Schedule of Events ................................................................................................ 25 
 
List of Figures 
Figure 1 Study Flow Chart .................................................................................................. 24 
 
 
 


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1. PROTOCOL SYNOPSIS (VERSION 3.0)  
Title 
A Pilot Study to Evaluate the Survival of Transplanted 
Staphylococcus Hominis A9 on the Skin of Adults with Moderate-to-
Severe Atopic Dermatitis 
Short Title 
Targeted Investigation of Microbiome Elimination (TIME-1) 
Clinical Phase 
Phase 1 
 
Number of Sites 
1 Clinical Site in the United States 
IND Sponsor/Number   
NIAID / IND # 17286 
Study Objectives 
Primary Objective 
To assess the duration of survival of coagulase negative 
staphylococcal species (CoNS) as measured by colony-forming 
unit (CFU) during a maximum of 24 days after the application of 
Staphylococcus hominis (S. hominis) A9 on the lesional ventral arm 
skin of Atopic Dermatitis (AD) participants positive for 
Staphylococcus aureus (S. aureus) (AD SA+). 
Secondary Objectives 
1. To assess the duration of survival of CoNS as measured by 
CFU during a maximum of 24 days after the application of 
S. hominis A9 on non-lesional ventral arm skin of AD 
participants positive for S. aureus (AD SA+). 
2. To assess the safety of S. hominis A9 or placebo 
application, as determined by the count of serious and non-
serious treatment- emergent adverse events (AEs) during 
the time period of Day 0 to Day 31 per participant within 
each group. 
Exploratory Objectives 
1. To assess the association of S. hominis A9 estimated CFU 
on the skin of the ventral arm contralateral to the arm of 
application and face. 
2. To assess the duration of survival of CoNS as measured by 
CFU during a maximum of 24 days after the application of 
S. hominis A9 on lesional ventral arm skin of AD 
participants negative for S. aureus (AD SA-).  
3. To assess the duration of survival of CoNS as measured by 
CFU during a maximum of 24 days after the application of 
S. hominis A9 on non-lesional ventral arm skin of AD 
participants negative for S. aureus (AD SA-). 
4. To compare the duration of survival of CoNS as measured 
by CFU on lesional skin between AD SA+ vs AD SA- 
participants. 
5. To compare the duration of survival of CoNS as measured 
by CFU on non-lesional skin between AD SA+ vs AD SA- 
participants. 
6. To compare the duration of survival of CoNS as measured 
by CFU between lesional and non-lesional skin on AD SA+ 
participants. 
7. To compare the duration of survival of CoNS as measured 
by CFU between lesional and non-lesional skin on AD SA- 
participants. 


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8. To compare disease severity measures (local Eczema Area 
and Severity Index [L-EASI], SCORing Atopic Dermatitis 
[SCORAD] and local Pruritus Numerical Rating Scale 
[NRS]) between Day 0 and the last in-clinic visit among AD 
SA+ participants, and independently, among AD SA-. 
9. To determine the antibiotic sensitivity of the skin CoNS 
microbiome to penicillin G, tetracycline, and erythromycin 
before and after treatment with ShA9. 
Study Design 
This is a Phase 1 pilot, open label, single center trial designed to 
assess the kinetics of S. hominis A9 survival on the skin of adults 
with moderate-to-severe atopic dermatitis on the ventral arms who 
are culture positive or negative for S. aureus colonization. 
Primary Endpoint 
The duration of survival, measured as the time needed for CoNS 
CFU to drop below baseline density measured before application of 
ShA9 + 100 CFU/cm2 on the lesional ventral arm skin of AD 
participants positive for S. aureus (AD SA+). 
Secondary Endpoints 
1. The duration of survival, measured as the time needed for 
CoNS CFU to drop below baseline density measured before 
application of ShA9 + 100 CFU/cm2 on non-lesional ventral 
arm skin of AD participants positive for S. aureus (AD SA+). 
2. The count of serious and non-serious treatment-emergent 
AEs per participant during the time period of Day 0 to Day 
31. 
Exploratory Endpoints 
1. Association of S. hominis A9 bacteria abundance estimated 
CFU as measured by % S. hominis A9 positive colonies by 
PCR x CoNS CFU while CoNS CFU is above baseline 
density measured before application of ShA9 + 100 
CFU/cm2 on the lesional and non-lesional, separately, 
ventral arm skin of AD participants between the arm treated 
with S. hominis A9 and the contralateral arm treated with 
placebo, and independently, with the participant’s face. 
2. The duration of survival, measured as the time needed for 
CoNS CFU to drop below baseline density measured before 
application of ShA9 + 100 CFU/cm2 on the lesional ventral 
arm skin of AD participants negative for S. aureus (AD SA-). 
3. The duration of survival, measured as the time needed for 
CoNS CFU to drop below baseline density measured before 
application of ShA9 + 100 CFU/cm2 on the non-lesional 
ventral arm skin of AD participants negative for S. aureus 
(AD SA-). 
4. Comparison of time needed for CoNS CFU to drop below 
baseline density measured before application of ShA9 + 
100 CFU/cm2 on the lesional and non-lesional, separately, 
ventral arm skin between AD participants positive (AD SA+) 
vs negative (AD SA-) for S. aureus. 
5. Comparison of time needed for CoNS CFU to drop below 
baseline density measured before application of ShA9 + 
100 CFU/cm2 on lesional vs non-lesional skin of AD 
participants positive (AD SA+) and negative (AD SA-) for S. 
aureus, separately. 


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6. Comparison of the L-EASI score of the ventral arms at Day 
0 vs the last in-clinic visit among AD SA+, and 
independently, among AD SA-.  
7. Comparison of the SCORAD score at Day 0 vs the last in-
clinic visit among AD SA+, and independently, among AD 
SA-.  
8. Comparison of the local Pruritus NRS of the ventral arms at 
Day 0 vs the last in-clinic visit among AD SA+, and 
independently, among AD SA-.  
9. Comparison of the proportion of antibiotic-sensitive CoNS to 
penicillin G, tetracycline, and erythromycin isolated from 
skin swabs at baseline and after ShA9 treatment, at the last 
swab collection from lesional skin. 
Accrual Objective 
This study will enroll a minimum of 20 adult participants (a 
minimum of 13 S. aureus positive and a minimum of 7 S. aureus 
negative), 18-80 years of age, with moderate-to-severe atopic 
dermatitis on the ventral arms. 
Study Duration 
This study will take approximately 18 months to complete 
participant recruitment, enrollment, and follow-up. An individual 
participant’s participation in this study will consist of up to 2 weeks 
of Screening, a 1-week pre-treatment phase, 1 day of treatment, 
and a maximum of 31 days for follow-up. Screened participants 
who meet all inclusion and exclusion criteria, including medication 
and therapy washouts, may complete Screening and Pre-treatment 
on the same day. 
Treatment Description  
(Investigational Products) 
Active (ShA9): 85% Phosphate-buffered saline solution (PBS) and 
15% Glycerol containing healthy donor-derived (allogeneic) 
commensal Staph species, S. hominis A9 (Manufactured and 
packaged by University of California – San Diego [UCSD]) applied 
once to the right or left ventral arm (wrist to upper arm)  
Placebo: 85% PBS and 15% Glycerol (Manufactured and packaged 
by UCSD) applied once to the contralateral ventral arm  
Inclusion Criteria 
Individuals who meet all of the following criteria are eligible for 
enrollment as study participants: 
1. Participant must be able to understand and provide 
informed consent. 
2. Male or female participants 18 to 80 years of age, inclusive 
at time of the Screening Visit  
3. Meet ADRN Standard Diagnostic Criteria (Appendix A) for 
active AD. 
4. At least 21 cm2 of lesional and 21 cm2 of non-lesional skin 
on both the right and left ventral arms. The required area 
(lesional or non-lesional) may be one contiguous area or 
may encompass multiple areas with a total cumulative area 
of 21 cm2. 
5. An Investigator Global Assessment (IGA) score, on the 
ventral arms of at least moderate severity 
6. Body surface area (BSA), as measured by Mostellar BSA 
Calculator, between 1.26 m2 and 2.25 m2.  


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7. If female of childbearing potential, must agree to remain 
abstinent (refrain from heterosexual intercourse) or use 
acceptable contraception (e.g., oral contraceptives, 
intrauterine device (IUD), barrier method with spermicide, 
surgically sterilized partner, Depo-Provera, Norplant, 
NuvaRing, or hormonal implants) for the duration of study 
participation. 
Exclusion Criteria 
Individuals who meet any of these criteria are not eligible for 
enrollment as study participants: 
1. Inability or unwillingness of a participant to give written 
informed consent or comply with study protocol. 
2. Pregnant or lactating females  
3. Active bacterial, viral, or fungal skin infections 
4. Any noticeable breaks or cracks in the skin on the target 
areas of investigational product application, including 
severely excoriated skin or skin with open or weeping 
wounds suggestive of an active infection or increased 
susceptibility to infection. 
5. Sensitivity to or difficulty tolerating Dove fragrance-free bar 
soap, Cetaphil® lotion, alcohol-based cleaners, glycerol, or 
soy products. 
6. Participants with Netherton’s syndrome or other 
genodermatoses that result in a defective epidermal barrier. 
7. Any participant who is immunocompromised (e.g., history of 
lymphoma, Human Immunodeficiency Virus (HIV)/Acquired 
Immunodeficiency Syndrome (AIDS), Wiskott-Aldrich 
Syndrome), has an immune system disorder (e.g., 
autoimmune disease), or is using a systemic 
immunosuppressant (e.g., systemic corticosteroids, 
cyclosporine, methotrexate) 
8. Any participant with current malignant disease (with the 
exception of non-melanoma skin cancer in an area not 
affected by treatment) 
9. Participants with a history of psychiatric disease or history 
of alcohol or drug abuse that would interfere with the ability 
to comply with the study protocol. 
10. Past or current medical problems or findings from physical 
examination or laboratory testing that are not listed above, 
which, in the opinion of the investigator, may pose 
additional risks from participation in the study, may interfere 
with the participant’s ability to comply with study 
requirements or that may impact the quality or interpretation 
of the data obtained from the study 
11. Ongoing participation in another investigational trial or use 
of investigational drugs within 8 weeks, or 5 half-lives (if 
known), whichever is longer, of the Screening Visit 
12. Treatment with non-steroid systemic immunosuppressant 
within 6 months of the Screening Visit 
13. Treatment with Dupilumab within 16 weeks of the Screening 
Visit 


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14. Treatment with oral or injectable therapy for AD (excluding 
oral steroids) within 5 half-lives (if known) or 16 weeks 
before the Screening Visit, whichever is longer.  
15. Participants with close contacts (e.g., spouse, children, or 
members in the same household) that have severe barrier 
defects or are immunocompromised. 
16. Use of topical (including steroids and calcineurin inhibitors) 
AD treatments on the ventral arms or face within 7 days of 
the Treatment Visit; Use of topical steroids on areas outside 
of where investigational product is to be applied or 
swabbing is to be performed may be permitted, per 
investigator discretion. 
17. Treatment with prescription moisturizers classified as 
medical device (e.g., Atopiclair®, MimyX®, Epiceram®, 
etc.) on the ventral arms or face within 7 days of the 
Treatment Visit; Use on areas outside of where 
investigational product is to be applied or swabbing is to be 
performed is permitted. 
18. Use of any oral or topical antibiotic within 7 days of the 
Treatment Visit 
19. Participants who have taken a bleach bath within 7 days of 
the Treatment Visit  
20. Use of any oral steroid therapies within 28 days of the 
Treatment Visit 
21. Any phototherapy for skin disease (such as narrow band 
ultraviolet B [NBUVB], ultraviolet B [UVB], ultraviolet A1 
[UVA1], psoralen + UVA [PUVA]) or regular use (more than 
2 visits per week) of a tanning bed within 28 days of the 
Treatment Visit 
Study Stopping Rules 
Study enrollment will be suspended pending Data and Safety 
Monitoring Board (DSMB) expedited review of all pertinent data if 
any of the following occur: 
1. A single participant experiences any Serious Adverse Event 
(SAE) for which there is a reasonable possibility that the 
investigational product caused the SAE. 
2. The development of any severe (Grade 3) AE for which 
attribution is defined as related or possibly related  
 
 


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2. INTRODUCTION 
This statistical analysis plan encompasses pre-determined analyses corresponding to the study 
objectives delineated in the protocol. This Phase 1 pilot, open-label, single-center trial is designed to 
evaluate the kinetics of S. hominis A9 survival on the skin of adults with moderate-to-severe atopic 
dermatitis on the ventral arms, regardless of S. aureus colonization status. The statistical analysis plan 
(SAP) offers comprehensive information to facilitate the execution of statistical analysis (interim 
analysis and final analysis) and reporting of the study data for use in the final Clinical Study Report 
(CSR). This SAP describes the populations that will be analyzed; the participant characteristic 
parameters, the efficacy parameters, and the safety parameters that will be evaluated; and details of 
the specific statistical methods that will be used. If additional analyses are required to supplement the 
planned analyses described in this SAP, they may be completed and will be identified in the CSR. 
Table, figure, and listing specifications are provided in separate documents. 
 
 


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3. GENERAL ANALYSIS AND REPORTING CONVENTIONS 
The following analyses and reporting conventions will be used: 
 
• 
Categorical variables will be summarized using counts (n) and percentages (%) and will be 
presented in the form “n (%).” Percentages will be rounded to one decimal place. 
• 
Numeric variables will be summarized using n, mean, standard deviation (SD), median, 
minimum (min), maximum (max). For data that follow a lognormal distribution (i.e., CoNS CFU), 
geometric mean and geometric SD will be reported instead of mean and SD.  
o The min/max will be reported at the same level of significance as original data. 
o The mean and median will be reported at one more significant digit than the precision of 
the data, and SD will be reported at two more significant digits than the precision of the 
data. 
o The median will be reported as the average of the two middle numbers if the dataset 
contains an even number of observations. 
• 
Test statistics including t and z test statistics will be reported to two decimal places.  
• 
P-values will be reported to three decimal places if greater than or equal to 0.001.  If less than 
0.001, the value will be reported as “<0.001.”  A p-value greater than 0.999 will be reported as 
“>0.999”. 
 
If departures from these general conventions are present in the specific evaluations section of this 
SAP, then those conventions will take precedence over these general conventions.  
 
 


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4. ANALYSIS SAMPLES 
4.1 Safety Sample 
The safety sample, which will be used for all safety analyses, will include all participants who are 
enrolled and receive any amount of S. hominis A9/placebo. For analysis, S. aureus CFU results from 
swabs obtained prior to treatment application during the Treatment Visit (Day 0) will be used to define 
SA+ vs SA- status of participants. 
 
4.2 Modified Intent-to-Treat (mITT) Sample 
The modified intent-to-treat sample will include all participants who are enrolled and receive any 
amount of S. hominis A9/placebo. For analysis, S. aureus CFU results from swabs obtained prior to 
treatment application during the Treatment Visit (Day 0) will be used to define SA+ vs SA- status of 
participants. 
 
4.3 Per-Protocol (PP) Sample 
The per-protocol sample will include all mITT participants who have no major protocol deviations in the 
category study medications. Specifically, participants who have no major protocol deviations of the 
following type: (1) administered study product not approved for distribution (i.e., expired, opened, 
contaminated, etc.) or (2) the wrong treatment dose (dose does not equal 1 mL or 6 (six) pumps total 
of placebo or treatment spray per arm). For analysis, S. aureus CFU results from swabs obtained prior 
to treatment application during the Treatment Visit (Day 0) will be used to define SA+ vs SA- status of 
participants. 
 
4.4 Interim Analysis Sample 
The interim analysis sample will include all participants who meet the following criteria: 
 
• 
Positive for S. aureus colonization prior to treatment application at the Treatment Visit (Day 0). 
• 
Application of the ShA9 and placebo treatments. 
 
 
 
 


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5. STUDY PARTICIPANTS 
5.1 Disposition of Participants  
The disposition (i.e., Recruited, Consented, Screened, Eligible, Enrolled, Completed, Discontinued) of 
all participants will be summarized in tables. 
 
The numbers and percentages of participants enrolled, the numbers and percentages of participants 
who completed the End of Study Phone Visit (Day 31), as well as reasons for early termination from 
the study will be presented across S. aureus positive/S. aureus negative status. Moreover, the study 
will stratify by arm dominance, with the dominant arm being the right arm for right-handed participants, 
and the left arm for left-handed participants. 
 
For participants who fail inclusion/exclusion criteria, failed criteria will be listed by participant. 
 
5.2 Demographic and Other Baseline Characteristics  
Summary descriptive statistics for demographic characteristics will be reported using conventions from 
section 3.0. Demographic characteristics collected at the Screening Visit (Day -14 to Day -7) that will 
be summarized include age (years), sex, ethnicity, race and Body Mass Index (BMI). 
 
A listing for vital signs will be reported. Vital signs and growth parameters are collected during all visits 
except the End of Study Phone Visit (Day 31) and include systolic blood pressure (mmHg), diastolic 
blood pressure (mmHg), heart rate (bpm), respiratory rate (respirations/min), temperature (F), height 
(in), weight (lbs), and body surface area (m2). 
 
Summary descriptive statistics for atopic dermatitis will be reported across arm dominance. Atopic 
dermatitis severity will be assessed during all visits except the End of Study Phone Visit (Day 31). 
Atopic dermatitis will be summarized by investigator’s global assessment (IGA), local eczema area 
and severity index (EASI) score on ventral arms, scoring atopic dermatitis (SCORAD) score, and local 
pruritus numerical rating scale (NRS) of ventral arms. 
 
5.3 Medical History  
Medical history is collected during the Screening (Day -14 to Day -7), Pre-Treatment (Day -7), and 
Treatment (Day 0) visits and will be reported in a listing. An abbreviated medical history will be 
collected during the Pre-Treatment (Day -7) and Treatment (Day 0) visits.  
 
6. STUDY OPERATIONS 
6.1 Protocol Deviations 
Protocol deviations will be listed with information such as type of deviation, severity of the deviation 
(major or non-major), date of occurrence, and the reason for the deviation. Protocol deviations will be 
summarized by type of deviation. 
 
6.2 Treatment Adherence 
Treatment adherence is not applicable to this study as this is a single application administered in clinic.  
 
 
 
 
 
 
 


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7. ENDPOINT EVALUATION 
7.1 Overview of Efficacy Analysis Methods 
7.1.1 Handling of Dropouts or Missing Data 
All efforts will be made via the querying and monitoring phases to avoid missing data. In general, 
missing data will not be imputed. 
 
The data will not be imputed for the primary endpoint. For more details, see Section 7.3 
 
7.1.2 Multicenter Studies 
Study participants will be recruited from 1 study site, University of California San Diego. 
 
7.1.3 Assessment Time Windows 
Allowable visit windows for all scheduled visits are provided in Section 14.2. All data will be included in 
analyses, regardless of time of assessment. 
 
Unscheduled visits may also occur throughout the study. Data from unscheduled visits will be included 
in listings but will generally not be included in tabular or graphical summaries. The one exception is if 
the unscheduled visit occurs during the allowable visit window of a missed visit, any available data 
from the unscheduled visit will be included in the summary of the missed visit data. 
 
7.2 Reporting of Swab Results 
Prior to the application of the ShA9 and placebo treatment, skin swabs will be collected from the 
lesional and non-lesional sites on the ventral arms and one non-lesional site on the participant’s face. 
After IP administration to each arm, additional swabs will be collected from the arms 15 minutes and 1, 
2, 4, and 6 hours after application. Participants will return to the clinic 24 hours after application (24 
Hour Visit) and on Days 3, 10, 17, and 24 for the collection of skin swabs from the identified target 
sites, as needed. However, after completion of the Day 3 Visit, a participant will not be required to 
complete the remaining follow up clinic visits (Day 10 through Day 24) once their lesional swabs from 
both arms (right and left) are CoNS negative, defined as below baseline density measured before 
application of ShA9 + 100 CFU/cm2. The collected swabs will be used to assess the following: 
 
1. S. aureus (CFU/cm2) 
2. Coagulase negative staphylococcal species (CoNS) (CFU/cm2) 
3. S. hominis A9 (CFU/cm2) 
4. Combined S. hominis A9 (CFU/cm2) 
5. Combined bacteria DNA (CFU/cm2) 
6. Antibiotic-sensitive CoNS 
 
S. aureus will be measured by counting CFUs on the Baird-Parker agar plate. Coagulase negative 
staphylococcal species (CoNS) will be measured by counting CFUs on the mannitol salt agar with egg 
yolk plate. CFUs of S. hominis A9 will be estimated from total CoNS CFU × % of S. hominis A9 
positive colonies. Abundance of bacterial DNA from total staphylococcus (G-staph), S. aureus, S. 
hominis, and Sh-lantibiotic will be quantified by qPCR. Antibiotic-sensitive CoNS will be estimated from 
[(CFU without antibiotic containing plate - CFU with antibiotic containing plate)/(CFU without antibiotic 
containing plate)]. 
 
Descriptive statistics and listings by participant of swab results will be presented. Swab results will be 
plotted to show the trajectory of results over time. Data will be plotted as a spaghetti plot where each 


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participant’s values will be plotted and connected by line segments, forming one line per participant. 
Plots will also indicate arm dominance per participant.  
 
7.3 Primary Objective 
The primary objective for the study is to assess duration of survival of coagulase negative 
staphylococcal species (CoNS) as measured by colony-forming unit (CFU) during a maximum of 24 
days after the application of S. hominis A9 on the lesional ventral arm skin of Atopic Dermatitis (AD) 
participants positive for S. aureus (AD SA+). 
 
7.3.1 Computation of the Primary Endpoint 
Time-to-CoNS elimination is defined as the time from Day 0 to the first occurrence of a value below 
baseline density measured before application of ShA9 + 100 CFU/cm2. The time to event for 
participants who did not reach CoNS CFU below baseline density measured before application of 
ShA9 + 100 CFU/cm2 during the study will be right-censored.  
 
7.3.2 Primary Analysis of the Primary Objective 
The primary endpoint for this pilot study is the time needed for CoNS CFU to drop below baseline 
density measured before application of ShA9 + 100 CFU/cm2 (time-to-CoNS elimination) in AD 
participants positive for S. aureus colonization. Time-to-CoNS elimination will be estimated using 
Kaplan Meier procedure.  
 
Median survival time along with 95% CI will be constructed based on a log-log transformed CI for the 
survivor function S(t). A graph of the Kaplan-Meier survival curve will be presented, and rates at fixed 
days will be derived (Therneau, et al., 2000). Furthermore, estimation of the hazard rate and 95% 
confidence interval will be produced using a parametric survival model.  
 
The following SAS code will be used to execute this plan:  
 
PROC LIFETEST data=data method=KM conftype=loglog plots=(survival(atrisk), hazard); 
    title 'Kaplan-Meier plot of CONS CFU Below Baseline (Log-Log)'; 
    TIME TIME*EVENT(0); 
run; 
 
The primary analysis will be conducted on the modified intent to treat sample and repeated on the per-
protocol sample if it differs from the modified intent to treat sample. 
 
7.3.3 Supportive Analyses of the Primary Endpoint 
We propose fitting a nonlinear time-response regression to our data set, which represents the 
relationship between the concentration of CoNS (Colony Forming Units per cm2) and time. The 
analysis will be conducted in R, leveraging the package drc known for its robust functionality in the 


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area of dose-response analysis. Specifically, we will model our data using either a two-parameter or a 
three-parameter log-logistic regression model. 
 
The following R code will be used to execute this plan: 
 
# Load the necessary package 
library(drc) 
 
# Fit a two-parameter log-logistic model 
model_2par <- drm(CoNS_CFU_per_cm2 ~ time, data = data, fct = LL.2()) 
 
# Fit a three-parameter log-logistic model 
model_3par <- drm(CoNS_CFU_per_cm2 ~ time, data = data, fct = LL.3()) 
 
# Compare the AIC values of the two models 
aic_2par <- AIC(model_2par) 
aic_3par <- AIC(model_3par) 
 
# Select the model with the lowest AIC 
best_model <- ifelse(aic_2par < aic_3par, "Two-parameter model", "Three-parameter model") 
 
# Compute the ED50 (or T50) from the selected model 
if (best_model == "Two-parameter model") { 
  ED50 <- ED(model_2par, 50) 
} else { 
  ED50 <- ED(model_3par, 50) 
} 
 
After conducting the model fits, we will evaluate the Akaike Information Criterion (AIC) for each model 
to compare their fit. The model yielding the lowest AIC will be selected as the primary model. From this 
primary model, we will determine the ED50 (or T50) - the time at which half of the maximum CoNS 
CFU/cm2 is achieved. This approach follows the methodology outlined by Ritz et al., 2015. 
Adjustments or additional steps may be needed depending on the specifics of the data set and the 
research question at hand. 
 
The supportive primary analysis will be conducted on the modified intent to treat sample and repeated 
on the per-protocol sample if it differs from the modified intent to treat sample. 
 
7.4 Secondary Objectives 
The secondary objectives are stated below: 
 
1. To assess the duration of survival of CoNS as measured by CFU during a maximum of 24 days 
after the application of S. hominis A9 on non-lesional ventral arm skin of AD participants positive 
for S. aureus (AD SA+).  
2. To assess the safety of S. hominis A9 or placebo application, as determined by the count of 
serious and non-serious treatment-emergent adverse events (AEs) during the time period of 
Day 0 to Day 31 per participant within each group. 
 
7.4.1 Secondary Endpoints 
1. The duration of survival, measured as the time needed for CoNS CFU to drop below baseline 
density measured before application of ShA9 + 100 CFU/cm2 on non-lesional ventral arm skin of 
AD participants positive for S. aureus (AD SA+). 
2. The count of serious and non-serious treatment-emergent AEs per participant during the time 
period of Day 0 to Day 31. 


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7.4.2 Analyses of Secondary Endpoints 
Secondary endpoint #1 will be analyzed using the same approach as described for the primary analysis 
of the primary endpoint (see Section 7.3.2). Median survival time along with 95% CI will be constructed 
based on a log-log transformed CI for the survivor function S(t). A graph of the Kaplan-Meier survival 
curve will be presented, and rates at fixed days will be derived (Therneau, et al., 2000). Furthermore, 
estimation of the hazard rate and 95% CI will be produced using a parametric survival model. 
 
The summary of all treatment-emergent adverse events and treatment-emergent serious adverse 
events for secondary endpoint #2 will be provided by system organ class, high-level term, and preferred 
term in descending order of frequency. 
 
Secondary endpoint #1 will be run on the modified intent to treat sample and re-run using the per-
protocol sample if this differs from the former. Secondary endpoint #2 will be run on the safety sample. 
 
7.5 Exploratory Objectives 
The exploratory objectives are stated below: 
 
1. To assess the association of S. hominis A9 estimated CFU on the skin of the ventral arm 
contralateral to the arm of application and face. 
2. To assess the duration of survival of CoNS as measured by CFU during a maximum of 24 days 
after the application of S. hominis A9 on lesional ventral arm skin of AD participants negative for 
S. aureus (AD SA-).  
3. To assess the duration of survival of CoNS as measured by CFU during a maximum of 24 days 
after the application of S. hominis A9 on non-lesional ventral arm skin of AD participants 
negative for S. aureus (AD SA-). 
4. To compare the duration of survival of CoNS as measured by CFU on lesional skin between AD 
SA+ vs AD SA- participants. 
5. To compare the duration of survival of CoNS as measured by CFU on non-lesional skin 
between AD SA+ vs AD SA- participants. 
6. To compare the duration of survival of CoNS as measured by CFU between lesional and non-
lesional skin on AD SA+ participants. 
7. To compare the duration of survival of CoNS as measured by CFU between lesional and non-
lesional skin on AD SA- participants. 
8. To compare disease severity measures (local Eczema Area and Severity Index [L-EASI], 
SCORing Atopic Dermatitis [SCORAD] and local Pruritus Numerical Rating Scale [NRS]) 
between Day 0 and the last in-clinic visit among AD SA+ participants, and independently, 
among AD SA-. 
9. To determine the antibiotic sensitivity of the skin CoNS microbiome to penicillin G, tetracycline, 
and erythromycin before and after treatment with ShA9. 
 
7.5.1 Exploratory Endpoints 
1. Association of S. hominis A9 bacteria abundance estimated CFU as measured by % S. hominis 
A9 positive colonies by PCR x CoNS CFU while CoNS CFU is above baseline density 
measured before application of ShA9 + 100 CFU/cm2 on the lesional and non-lesional, 
separately, ventral arm skin of AD participants between the arm treated with S. hominis A9 and 
the contralateral arm treated with placebo, and independently, with the participant’s face. 
2. The duration of survival, measured as the time needed for CoNS CFU to drop below baseline 
density measured before application of ShA9 + 100 CFU/cm2 on the lesional ventral arm skin of 
AD participants negative for S. aureus (AD SA-). 
3. The duration of survival, measured as the time needed for CoNS CFU to drop below baseline 
density measured before application of ShA9 + 100 CFU/cm2 on the non-lesional ventral arm 
skin of AD participants negative for S. aureus (AD SA-). 


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4. Comparison of time needed for CoNS CFU to drop below baseline density measured before 
application of ShA9 + 100 CFU/cm2 on the lesional and non-lesional, separately, ventral arm 
skin between AD participants positive (AD SA+) vs negative (AD SA-) for S. aureus. 
5. Comparison of time needed for CoNS CFU to drop below baseline density measured before 
application of ShA9 + 100 CFU/cm2 on lesional vs non-lesional skin of AD participants positive 
(AD SA+) and negative (AD SA-) for S. aureus, separately. 
6. Comparison of the L-EASI score of the ventral arms at Day 0 vs the last in-clinic visit among AD 
SA+, and independently, among AD SA-.  
7. Comparison of the SCORAD score at Day 0 vs the last in-clinic visit among AD SA+, and 
independently, among AD SA-. 
8. Comparison of the local Pruritus NRS of the ventral arms at Day 0 vs the last in-clinic visit 
among AD SA+, and independently, among AD SA-.  
9. Comparison of the proportion of antibiotic-sensitive CoNS to penicillin G, tetracycline, and 
erythromycin isolated from skin swabs at baseline and after ShA9 treatment, at the last swab 
collection from lesional skin. 
 
7.5.2 Analyses of the Exploratory Endpoints 
Repeated measures correlation will be calculated for determining the common within-individual 
association for paired (S. hominis treated arm and contralateral arm treated with placebo, and 
independently, with the participant’s face) for S. hominis A9 bacteria abundance estimated CFU as 
measured by % S. hominis A9 positive colonies by PCR x CoNS CFU assessed on two or more days. 
The correlation coefficient, the 95% confidence interval, and the p-value will be reported. Log 
transformation will likely be needed for the outcome of interest to produce a less skewed distribution 
(Bland and Altman, 1995).  
 
Analyses of disease severity (L-EASI, SCORAD, and local Pruritus NRS) change between Day 0 vs 
the last in-clinic visit among AD SA+, and independently, among AD SA- will be conducted using a 
repeated measures mixed-effects model with an unstructured covariance matrix. The model will 
estimate the pre-intervention vs. post-intervention least squares mean ± standard error and the 
difference reported with a 95% confidence interval and p-value. 
 
Analyses to investigate if ShA9 application can change CoNS antibiotic sensitivity to penicillin G, 
tetracycline, and erythromycin will be conducted on all participants (AD SA+ and AD SA-) by 
comparing the proportion of antibiotic-sensitive CoNS [(CFU without antibiotic containing plate - CFU 
with antibiotic containing plate) / CFU without antibiotic containing plate] before and after treatment. 
We will model the difference in proportions of antibiotic-sensitive CoNS between swabs taken at 
baseline and swabs taken at the last Follow-Up Visit. 
 
The exploratory analyses will be conducted on the modified intent-to-treat sample, and due to the 
exploratory nature of this endpoint analysis, no adjustments will be made for multiple comparisons. 
 
8. SAFETY EVALUATION 
8.1 Overview of Safety Analysis Methods 
All safety analyses will be carried out using the safety sample. Missing safety information will not be 
imputed. Listings will be prepared for all safety measurements. All listings will be sorted in order of S. 


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aureus positive/S. aureus negative status, participant identifier, and the time of assessment (e.g., visit, 
time, and/or event). 
 
8.2 Adverse Events 
All AEs will be classified by system organ class (SOC) and preferred term, according to a standardized 
thesaurus (Medical Dictionary for Regulatory Activities [MedDRA] version 25.0). 
Each AE is entered on the eCRF once at the highest severity. As such, no additional data 
manipulation is needed to identify events. 
 
AEs will be collected from the time of consent until the participant completes study participation or until 
30 days after he/she prematurely withdraws (without withdrawing consent) or is withdrawn from the 
study. Treatment-emergent AEs are defined as new AEs or an increase in grade of an AE after 
investigational product application (Day 0).  
 
An overall summary table will be developed to report the number of events and the number and 
percentage of participants having at least one event in the following categories:  
 
• 
AEs 
• 
AEs indicated as serious 
• 
AEs with an outcome of death 
• 
AEs that were reported as being related to the study treatment 
• 
AEs reported by maximum severity 
 
In addition, AEs classified by MedDRA SOC, and preferred term will be summarized by arm 
dominance and overall, for each of the following: 
 
• 
All AEs 
• 
AEs by maximum severity 
• 
AEs by relationship to study treatment 
 
Summary tables will present the total number of events as well as the number and percentage of 
participants experiencing the events. If a participant experiences the same AE on multiple occasions, 
the event will be counted once for each occurrence when reporting the number of AEs. When 
reporting the number of participants experiencing the events, a participant will only be counted once if 
he/she experiences an event within the particular SOC or preferred term.  
 
8.3 Deaths and Serious Adverse Events 
Serious adverse events (SAEs) will be listed and summarized in the same manner described in 
Section 8.2. Separate displays listing and summarizing death, including time to death and cause of 
death, will also be created. 
 
8.4 Vital Signs, Physical Findings, and Other Observations Related to Safety 
8.4.1 Vital Signs 
Data listings sorted by S. aureus positive/S. aureus negative participant status, vital sign parameter, 
and the time of assessment will be provided for vital sign measurements. Vital signs include systolic 
blood pressure (mmHg), diastolic blood pressure (mmHg), heart rate (bpm), respiratory rate 
(respirations/min), temperature (F), height (in), weight (lbs), and body surface area (m2). 
 
8.4.2 Pregnancy Assessment  
Pregnancy tests (urine) will be performed on female participants of child-bearing potential who do not 
self-report as pregnant at the Screening (Day -14 to Day -7) and Treatment (Day 0) visits. If a female 
participant becomes pregnant prior to the Treatment Visit (Day 0), the participant will no longer be 


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eligible to participate in the study and will be withdrawn from all future study activities listed in the 
Schedule of Events.  
 
At all other study visits, female participants of child-bearing potential will be asked if they are pregnant. 
If a pregnancy occurs, the pregnant participant will continue all study activities listed in the Schedule of 
Events. All pregnancies identified during the study shall be followed to conclusion and reported in a 
listing by participant. If pregnancy complications result in a congenital abnormality, birth defect, 
miscarriage, or medically indicated abortion, an SAE shall be reported.  
 
9. OTHER ANALYSES 
9.1 Use of Medications 
Medications will be coded according to the World Health Organization (WHO) Drug Dictionary (version 
2017.01). Medications reported on the eCRF will be categorized for analysis as prior, concomitant, or 
after study treatment by comparing the medication start and stop dates with the study treatment date. 
During study participation, participants are permitted to use topical steroids per discretion of the 
investigator on areas other than the ventral arms where treatment is applied and the designated 
swabbing area on the face. 
 
The number and percentage of participants receiving prior, concomitant, and post-treatment 
medications will be presented overall and by medication class. When reporting the number of 
participants receiving the medication, a participant will only be counted once if they ever received the 
medication within the medication class. Percentages will be based on the number of participants in the 
analysis population. 
 
10. INTERIM ANALYSES AND DATA MONITORING 
10.1 Interim Safety Reporting to the Data and Safety Monitoring Board 
The progress of the study will be monitored by the Data and Safety Monitoring Board (DSMB). The 
National Institute of Allergy and Infectious Diseases (NIAID) Allergy-Asthma [Alpha] DSMB will be 
chartered to review safety data and to make recommendations regarding continuation, termination, or 
modification of the study. The DSMB will formally review the safety data at least yearly.  
 
In addition, safety data will be reviewed by the DSMB when an event occurs that is of sufficient concern 
to the NIAID medical monitor or protocol chair to warrant review, or when an event occurs that could 
contribute to a predefined stopping rule specified in the protocol. However, no formal interim analysis of 
safety data will be conducted.  
 
10.2 Interim Analysis of Primary Endpoint 
An interim analysis will be performed to inform the development of upcoming trials, especially with 
regards to their design and power/sample size considerations. This analysis will be performed when 
complete data for 50% of AD participants positive for S. aureus colonization becomes available. For 
analysis, S. aureus CFU results from swabs obtained prior to treatment application during the 
Treatment Visit will be used to define SA+ vs SA- status of participants. Duration of survival of 
coagulase negative staphylococcal species (CoNS) as measured by colony-forming unit (CFU) will be 
evaluated. Enrollment will not be impacted or paused while the interim analysis is being performed. 
 
Time-to-CoNS elimination will be estimated using Kaplan Meier techniques. Median survival time along 
with 95% CI will be constructed based on a log-log transformed CI for the survivor function S(t). A 
graph of the Kaplan-Meier survival curve will be presented, and rates at fixed days will be derived 
(Therneau, et al., 2000). Furthermore, estimation of the hazard rate and 95% confidence interval will be 
produced using a parametric survival model. Summary statistics will be reported, subject-level data will 
not be presented for the interim analysis as only tabular group level (by arm dominance) summaries will 
be included. Furthermore, since all participants involved in the interim analysis will have completed the 


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study, there will be no risk in biasing site staff concerning AE reporting for these participants. AE data 
will not be analyzed in any fashion for the interim analysis. Blinding as to which arm of each participant 
received active treatment will be maintained for all fully blinded study staff, according to the guidelines 
defined in the Randomization Plan. Tabular group level (by arm dominance) summaries will be reported 
to the protocol chair Dr. Richard Gallo, the ADRN principal investigator Dr. Donald Leung, and staff at 
the NIAID and Rho. 
 
11. SAMPLE SIZE CONSIDERATIONS  
The proposed sample size for this study is 20 participants (13 participants in the S. aureus positive and 
7 in the S. aureus negative arm (2:1 randomization)). The primary objective of the analysis is to 
estimate the duration of survival of CoNS as measured by CFU during a maximum of 24 days after the 
application of S. hominis A9 on the lesional ventral arm skin of AD participants positive for S. aureus 
(AD SA+).  
 
The sample size was derived from ADRN-08: Targeted Microbiome Transplant in Atopic Dermatitis 
where, using a parametric survival model with exponential distribution, we estimated a hazard rate (λ) 
of approximately 0.09. This estimate was derived from a follow-up time of 11 days (shorter study) with 
35 participants where nine (25%) were right-censored at trial end. The goal for this trial is to estimate 
the hazard rate and its 95% confidence interval with precision for the width of the confidence interval as 
0.20. The 13 participants (9 events) proposed for the primary analysis were derived using these 
previous data as a starting point but considering more conservative estimates with a hazard rate of 0.15 
and 30% right censoring, which produces a two-sided 95% CI width equal to 0.194. 
 
12. CHANGES TO THE ANALYSES PLANNED IN THE PROTOCOL 
A per-protocol (pp) analysis sample not originally defined in the protocol has been added to the 
statistical analysis plan. 
 
The planned analysis for secondary endpoint #1 described in Section 13.4.4 of the Protocol does not 
correctly describe an appropriate analysis plan for that endpoint. Section 7.4.2 of the SAP has been 
updated to correctly describe the intended planned analysis for secondary endpoint #1.  
 
 


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13. REFERENCES 
 
Bland, J.M., & Altman, D.G. (1995). Calculating correlation coefficients with repeated observations: Part 
1 - correlation within subjects. BMJ, 310, 446.  
 
Ritz, C., Baty, F., Streibig, J. C., Gerhard, D. (2015) Dose-Response Analysis Using R PLOS ONE, 
10(12), e0146021.  
 
Therneau Terry M., Grambsch Patricia M. (2000).  Modeling Survival Data: Extending the Cox Model. 
Springer, New York. ISBN 0-387-98784-3. 
 
 


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14. APPENDICES  
14.1 Study Flow Chart 
Figure 1 Study Flow Chart 
 
 
 
 
 
 


14.2 Schedule of Events 
Study Visit 
Recruitment1 
Screening2 
Pre-treatment3 
Treatment  
Follow-Up Visits4 
End of Study 
Phone-Visit 
Unscheduled 
Visit5 
24 Hour 
Day 3 
Day 10 
Day 17 
Day 24 
Day (D), Visit Window 
 
Day -14 
to  
Day -7 
Day -7 
± 
1 Day  
Day 0 
Day 1 
Day 3 ± 
1 Day 
Day 10 ± 
1 Day 
Day 17 ± 
2 Days 
Day 24 
± 
2 Days 
Day 31 ±  
7 Days 
 
Study Assessments 
Recruitment Script 
X 
 
 
 
 
 
 
 
 
 
 
Informed Consent 
 
X 
 
 
 
 
 
 
 
 
 
Demographics 
 
X 
 
 
 
 
 
 
 
 
 
Medical History 
 
X 
X6 
X6 
 
 
 
 
 
 
 
Physical Exam 
 
X 
X 
X 
 
 
 
 
 
 
X 
AD Severity Assessment7 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
Pregnancy Status & Test8 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Concomitant Medications 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Vital Signs & Growth Parameters9 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
Skin Swab Collection10 
 
 
X11 
X12,13 
X 
X  
X 
X  
X  
  
X14 
AE Assessment 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Distribute Dove 
 
 
X 
 
 
 
 
 
 
 
 
Distribute Cetaphil 
 
 
X 
 
 
 
 
 
 
 
 
Randomization 
 
 
 
X 
 
 
 
 
 
 
 
ShA9 and Placebo Application 
 
 
 
X 
 
 
 
 
 
 
 
Distribute Handcard 
 
 
X 
X 
 
 
 
 
 
 
 
Blood Collection 
 
 
 
 
 
 
 
 
 
 
X14 


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Table 1 Schedule of Events 
1. Participants may be recruited by phone or during an in-person visit.  
2. Assessment of full inclusion and exclusion criteria will occur during the Screening Visit, after participants have consented to study participation. 
3. Participants who meet full inclusion and exclusion criteria including medication and therapy washouts, may complete the Screening and Pre-
treatment Visits on the same day. The following assessments will not be repeated, if the Pre-treatment Visit is conducted on the same day as 
Screening: medical history, physical exam, AD severity assessment, concomitant medications, pregnancy test, and vitals.   
4. Upon completion of the Day 3 Follow-Up Visit, a participant will be notified prior to their next scheduled follow-up visit if they need to return to 
the clinic. A participant will not be required to complete the remaining follow up clinic visits (Day 10 through Day 24) if the lesional swabs from 
both arms (right and left) collected at their most recent prior visit, have CoNS CFU below baseline density measured before application of 
ShA9 + 100 CFU/cm2.   
5. If disease activity increases, participants experience signs and symptoms as described on the instructional handcard, or other concerns arise 
between regularly scheduled visits, participants may be asked to return to the study site for an Unscheduled Visit. 
6. An abbreviated Medical History will be collected.  
7. AD disease severity will be assessed using the Investigator Global Assessment (Screening only), local Eczema Area and Severity Index (L-
EASI) on the arms, SCORing Atopic Dermatitis (SCORAD), and local Pruritus Numerical Rating Scale (NRS). 
8. A urine pregnancy test will be completed for all female participants of childbearing potential who do not self-report as pregnant at the 
Screening and Treatment Visits. 
9. Vital signs will include temperature, heart rate, respiration rate, systolic blood pressure, and diastolic blood pressure. Growth parameters 
(height and weight) will only be collected at the Screening Visit.  
10. Skin swab collection from the participant’s ventral arms will occur during each visit, and skin swab collection from the participant’s face will 
begin at the Treatment Visit. 
11. Lesional and non-lesional sites will be identified on the participant’s ventral arms for skin swab collection. Digital photographs of the lesional 
and non-lesional swab sites will be taken prior to swab collection. Each photograph will include a ruler so the scale of the sites can be 
determined. These sites will be swabbed at subsequent visits. 
12. A non-lesional site will be identified on the participant’s face for skin swab collection. A digital photograph of the non-lesional swab site will be 
taken prior to swab collection. The photograph will include a ruler so the scale of the site can be determined. This site will be swabbed at 
subsequent visits. 
13. Participants will remain in clinic for up to 6 hours following the application of investigational product during the Treatment Visit. Additional skin 
swabs will be collected from the ventral arms at 15 minutes, 1, 2, 4, and 6 hours post application.  
14. Skin swabs and/or blood may be collected during an Unscheduled Visit per investigator discretion 

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PROTOCOL ADRN-13  
A PILOT STUDY TO EVALUATE THE SURVIVAL OF TRANSPLANTED STAPHYLOCOCCUS HOMINIS A9 ON THE SKIN 
OF ADULTS WITH MODERATE-TO-SEVERE ATOPIC DERMATITIS   
TARGETED INVESTIGATION OF MICROBIOME ELIMINATION (TIME-1)  
  
3.0 /28 JUL 2023  
  
INVESTIGATIONAL NEW DRUG (IND)# 17286   
National Clinical Trial (NCT)# NCT05177328  
IND SPONSOR: The National Institute of Allergy and Infectious Diseases (NIAID)   
NIAID Funding Mechanism: Grant 1 U01 AI152038-01 and Grant UM2AI117870  
  
Investigational Agent Manufacturer/Provider: University of California – San Diego  Consortium/Network: 
Atopic Dermatitis Research Network (ADRN)   
  
 
Confidentiality Statement  
The information contained within this document is not to be disclosed in any way without the prior permission of the Division of 
Allergy, Immunology, and Transplantation, National Institute of Allergy and Infectious Diseases, National Institutes of Health.  
  
  
PROTOCOL CHAIR- RICHARD GALLO, MD, PHD  
Distinguished Professor and Chairman,  
Department of Dermatology  
University of California – San Diego  
9500 Gilman Drive  
Mail code 0869  
La Jolla, CA 92093-0869  
Phone: 858-822-4608  
Fax: 858-822-6985  
E-mail: rgallo@health.ucsd.edu    
MEDICAL MONITOR- 
  
  
  
National Institute of Allergy and Infectious  
Diseases  
5601 Fishers Lane, Room 6B53  
Bethesda, MD 20892-9828, USA  
   
  
 
STATISTICAL AND CLINICAL COORDINATING   
CENTER (SACCC) PRINCIPAL INVESTIGATOR-  
  
   
Rho Federal Systems   
2635 East, NC-54  
Durham, NC 27713  
  
  
   
PROTOCOL CO-CHAIR- TISSA HATA, MD  
HS Professor of Dermatology,  
Department of Dermatology  
University of California – San Diego  
9500 Gilman Drive  
Mail code 0869  
La Jolla, CA 92093-0869  
Phone: 858-657-8390  
Fax: 858-657-8391  
E-mail: thata@health.ucsd.edu  
PROJECT MANAGER-
– 
  
  
 
  
  
National Institute of Allergy and Infectious  
Diseases  
5601 Fishers Lane, Room 6B51  
Bethesda, MD 20892-9828, USA Phone: 
  
  
   
REGULATORY OFFICER- 
  
  
  
  
National Institute of Allergy and  
Infectious Diseases  
5601 Fishers Lane, Room 7B46  
Bethesda, MD 20892-9828, USA Phone: 
   
  
    


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SITE INVESTIGATOR SIGNATURE PAGE  
Protocol Number: ADRN-13  
   
Version Number/Date: 3.0/ 28 Jul 2023  
   
Protocol Title:  A Pilot Study to Evaluate the Survival of Transplanted Staphylococcus Hominis A9 
on the Skin of Adults with Moderate-to-Severe Atopic Dermatitis    
   
IND Sponsor:  The National Institute of Allergy and Infectious Diseases (NIAID)       
Return Signed Form to:  
[The original signature page must be kept for your records.  Return an electronic PDF copy of the signed 
signature page (*as described below) to the 
 
 
  
  
I confirm that I have read the above protocol in the latest version.  I understand it, and I will work 
according to the principles of Good Clinical Practice (GCP) as described in the United States Code of 
Federal Regulations (CFR) – 45 CFR part 46 and 21 CFR parts 50, 56, 312, and 812 and in the  
International Conference for Harmonisation (ICH) document entitled Integrated Addendum to ICH 
E6(R1): Guideline for Good Clinical Practice E6(R2).  Further, I will conduct the study in keeping with local 
legal and regulatory requirements.  
As the site Principal Investigator, I agree to carry out the study by the criteria written in the protocol 
and understand that no changes can be made to this protocol without the written permission of the  
Institutional Review Board (IRB) and NIAID.  
[*The site Principal Investigator should print, sign, and date at the indicated location below.  A written 
signature is acceptable, and an electronic signature is acceptable in a pdf version of the form.]  
  
____________________________________      
Site Principal Investigator (Print)  
  
____________________________________                                      _________________  
Site Principal Investigator (Signature)                                                          Date  
 
 
 
 
 
 
 
 
 
 


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Protocol Synopsis  
Title  
A Pilot Study to Evaluate the Survival of Transplanted Staphylococcus Hominis A9 
on the Skin of Adults with Moderate-to-Severe Atopic Dermatitis  
Short Title  
Targeted Investigation of Microbiome Elimination (TIME-1)  
Clinical Phase  
Phase 1  
  
Number of Sites  
1 Clinical Site in the United States  
IND Sponsor/Number    
NIAID / IND # 17286  


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Study Objectives  
Primary Objective  
To assess the duration of survival of coagulase negative staphylococcal species 
(CoNS) as measured by colony-forming unit (CFU) during a maximum of 24 days 
after the application of Staphylococcus hominis (S. hominis) A9 on the lesional 
ventral arm skin of Atopic Dermatitis (AD) participants positive for 
Staphylococcus aureus (S. aureus) (AD SA+) Secondary Objectives  
1. To assess the duration of survival of CoNS as measured by CFU during a 
maximum of 24 days after the application of S. hominis A9 on nonlesional 
ventral arm skin of AD participants positive for S. aureus (AD SA+)  
2. To assess the safety of S. hominis A9 or placebo application, as 
determined by the count of serious and non-serious treatment- 
emergent adverse events (AEs) during the time period of Day 0 to Day 31 
per participant within each group Exploratory Objectives  
1. To assess the association of S. hominis A9 estimated CFU on the skin of 
the ventral arm contralateral to the arm of application and face  
2. To assess the duration of survival of CoNS as measured by CFU during a 
maximum of 24 days after the application of S. hominis A9 on lesional 
ventral arm skin of AD participants negative for S. aureus (AD SA-)   
3. To assess the duration of survival of CoNS as measured by CFU during a 
maximum of 24 days after the application of S. hominis A9 on 
nonlesional ventral arm skin of AD participants negative for S. aureus 
(AD SA-)  
4. To compare the duration of survival of CoNS as measured by CFU on 
lesional skin between AD SA+ vs AD SA- participants  
5. To compare the duration of survival of CoNS as measured by CFU on 
non-lesional skin between AD SA+ vs AD SA- participants  
6. To compare the duration of survival of CoNS as measured by CFU 
between lesional and non-lesional skin on AD SA+ participants  
 
 
7. To compare the duration of survival of CoNS as measured by CFU 
between lesional and non-lesional skin on AD SA- participants  
8. To compare disease severity measures (local Eczema Area and Severity 
Index [L-EASI], SCORing Atopic Dermatitis [SCORAD] and local Pruritus 
Numerical Rating Scale [NRS]) between Day 0 and the last in-clinic visit 
among AD SA+ participants, and independently, among AD SA-  
9. To determine the antibiotic sensitivity of the skin CoNS microbiome to 
penicillin G, tetracycline, and erythromycin before and after treatment 
with ShA9  


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Study Design  
This is a Phase 1 pilot, open label, single center trial designed to assess the 
kinetics of S. hominis A9 survival on the skin of adults with moderate-to-severe 
atopic dermatitis on the ventral arms who are culture positive or negative for S. 
aureus colonization.  
Primary Endpoint  
The duration of survival, measured as the time needed for CoNS CFU to drop 
below baseline density measured before application of ShA9 + 100 CFU/cm2 on 
the lesional ventral arm skin of AD participants positive for S. aureus (AD SA+)  
Secondary Endpoints  
1.  
The duration of survival, measured as the time needed for CoNS CFU to 
drop below baseline density measured before application of ShA9 + 100 
CFU/cm2 on non-lesional ventral arm skin of AD participants positive for  
S. aureus (AD SA+)  
 
2.  The count of serious and non-serious treatment-emergent AEs per 
participant during the time period of Day 0 to Day 31  
Exploratory Endpoints  
1.  Association of S. hominis A9 bacteria abundance estimated CFU as 
measured by % S. hominis A9 positive colonies by PCR x CoNS CFU while  
CoNS CFU is above baseline density measured before application of 
ShA9 + 100 CFU/cm2 on the lesional and non-lesional, separately, 
ventral arm skin of AD participants between the arm treated with S.  
hominis A9 and the contralateral arm treated with placebo, and 
independently, with the participant’s face  
 
2.  The duration of survival, measured as the time needed for CoNS CFU to 
drop below baseline density measured before application of ShA9 + 100  
CFU/cm2 on the lesional ventral arm skin of AD participants negative for 
S. aureus (AD SA-)  
 
3.  The duration of survival, measured as the time needed for CoNS CFU to 
drop below baseline density measured before application of ShA9 + 100 
CFU/cm2 on the non-lesional ventral arm skin of AD participants 
negative for S. aureus (AD SA-)  
 
4.  Comparison of time needed for CoNS CFU to drop below baseline 
density measured before application of ShA9 + 100 CFU/cm2 on the 
lesional and non-lesional, separately, ventral arm skin between AD 
participants positive (AD SA+) vs negative (AD SA-) for S. aureus  
 
5.  Comparison of time needed for CoNS CFU to drop below baseline 
density measured before application of ShA9 + 100 CFU/cm2 on lesional  
 


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vs non-lesional skin of AD participants positive (AD SA+) and negative (AD 
SA-) for S. aureus, separately  
6. Comparison of the L-EASI score of the ventral arms at Day 0 vs the last 
in-clinic visit among AD SA+, and independently, among AD SA-   
7. Comparison of the SCORAD score at Day 0 vs the last in-clinic visit 
among AD SA+, and independently, among AD SA-   
8. Comparison of the local Pruritus NRS of the ventral arms at Day 0 vs the 
last in-clinic visit among AD SA+, and independently, among AD SA-   
9. Comparison of the proportion of antibiotic-sensitive CoNS to penicillin 
G, tetracycline, and erythromycin isolated from skin swabs at baseline 
and after ShA9 treatment, at the last swab collection from lesional skin  
Accrual Objective  
This study will enroll a minimum of 20 adult participants (a minimum of 13 S.  
aureus positive and a minimum of 7 S. aureus negative), 18-80 years of age, with 
moderate-to-severe atopic dermatitis on the ventral arms.  
Study Duration  
This study will take approximately 18 months to complete participant 
recruitment, enrollment, and follow-up. An individual participant’s participation 
in this study will consist of up to 2 weeks of Screening, a 1-week pre-treatment 
phase, 1 day of treatment, and a maximum of 31 days for follow-up. Screened 
participants who meet all inclusion and exclusion criteria, including medication 
and therapy washouts, may complete Screening and Pre-treatment on the same 
day.  
Treatment Description   
(Investigational Products)  
Active (ShA9): 85% Phosphate-buffered saline solution (PBS) and 15% Glycerol 
containing healthy donor-derived (allogeneic) commensal Staph species, S. 
hominis A9 (Manufactured and packaged by University of California – San Diego  
[UCSD]) applied once to the right or left ventral arm (wrist to upper arm)   
Placebo: 85% PBS and 15% Glycerol (Manufactured and packaged by UCSD) 
applied once to the contralateral ventral arm   


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Inclusion Criteria  
Individuals who meet all of the following criteria are eligible for enrollment as 
study participants:  
1. Participant must be able to understand and provide informed consent  
2. Male or female participants 18 to 80 years of age, inclusive at time of 
the Screening Visit   
3. Meet ADRN Standard Diagnostic Criteria (Appendix A) for active AD  
4. At least 21 cm2 of lesional and 21 cm2 of non-lesional skin on both the 
right and left ventral arms. The required area (lesional or non-lesional) 
may be one contiguous area or may encompass multiple areas with a 
total cumulative area of 21 cm2.  
5. An Investigator Global Assessment (IGA) score, on the ventral arms of at 
least moderate severity  
6. Body surface area (BSA), as measured by Mostellar BSA Calculator, 
between 1.26 m2 and 2.25 m2   
7. If female of child bearing potential, must agree to remain abstinent  
(refrain from heterosexual intercourse) or use acceptable contraception  
 
 
(e.g., oral contraceptives, intrauterine device (IUD), barrier method with 
spermicide, surgically sterilized partner, Depo-Provera, Norplant, 
NuvaRing, or hormonal implants) for the duration of study participation  


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Exclusion Criteria  
Individuals who meet any of these criteria are not eligible for enrollment as study 
participants:  
1. Inability or unwillingness of a participant to give written informed 
consent or comply with study protocol  
2. Pregnant or lactating females   
3. Active bacterial, viral, or fungal skin infections  
4. Any noticeable breaks or cracks in the skin on the target areas of 
investigational product application, including severely excoriated skin or 
skin with open or weeping wounds suggestive of an active infection or 
increased susceptibility to infection  
5. Sensitivity to or difficulty tolerating Dove fragrance-free bar soap, 
Cetaphil® lotion, alcohol-based cleaners, glycerol, or soy products  
6. Participants with Netherton’s syndrome or other genodermatoses that 
result in a defective epidermal barrier  
7. Any participant who is immunocompromised (e.g., history of 
lymphoma, Human Immunodeficiency Virus (HIV)/Acquired  
Immunodeficiency Syndrome (AIDS), Wiskott-Aldrich Syndrome), has an 
immune system disorder (e.g., autoimmune disease), or is using a 
systemic immunosuppressant (e.g., systemic corticosteroids, 
cyclosporine, methotrexate)  
8. Any participant with current malignant disease (with the exception of 
non-melanoma skin cancer in an area not affected by treatment)  
9. Participants with a history of psychiatric disease or history of alcohol or 
drug abuse that would interfere with the ability to comply with the 
study protocol  
10. Past or current medical problems or findings from physical examination 
or laboratory testing that are not listed above, which, in the opinion of 
the investigator, may pose additional risks from participation in the 
study, may interfere with the participant’s ability to comply with study 
requirements or that may impact the quality or interpretation of the 
data obtained from the study  
11. Ongoing participation in another investigational trial or use of 
investigational drugs within 8 weeks, or 5 half-lives (if known), 
whichever is longer, of the Screening Visit  
12. Treatment with non-steroid systemic immunosuppressant within 6 
months of the Screening Visit  
13. Treatment with Dupilumab within 16 weeks of the Screening Visit  
14. Treatment with oral or injectable therapy for AD (excluding oral 
steroids) within 5 half-lives (if known) or 16 weeks before the Screening 
Visit, whichever is longer   
15. Participants with close contacts (e.g., spouse, children, or members in 
the same household) that have severe barrier defects or are 
immunocompromised  
 


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16. Use of topical (including steroids and calcineurin inhibitors) AD 
treatments on the ventral arms or face within 7 days of the Treatment 
Visit; Use of topical steroids on areas outside of where investigational 
product is to be applied or swabbing is to be performed may be 
permitted, per investigator discretion.  
17. Treatment with prescription moisturizers classified as medical device 
(e.g., Atopiclair®, MimyX®, Epiceram®, etc.) on the ventral arms or face 
within 7 days of the Treatment Visit; Use on areas outside of where 
investigational product is to be applied or swabbing is to be performed 
is permitted  
18. Use of any oral or topical antibiotic within 7 days of the Treatment Visit  
19. Participants who have taken a bleach bath within 7 days of the 
Treatment Visit   
20. Use of any oral steroid therapies within 28 days of the Treatment Visit  
21. Any phototherapy for skin disease (such as narrow band ultraviolet B  
[NBUVB], ultraviolet B [UVB], ultraviolet A1 [UVA1], psoralen + UVA 
[PUVA]) or regular use (more than 2 visits per week) of a tanning bed 
within 28 days of the Treatment Visit  
Study Stopping Rules  
Study enrollment will be suspended pending Data and Safety Monitoring Board 
(DSMB) expedited review of all pertinent data if any of the following occur:  
1. A single participant experiences any Serious Adverse Event (SAE) for 
which there is a reasonable possibility that the investigational product 
caused the SAE  
2. The development of any severe (Grade 3) AE for which attribution is 
defined as related or possibly related   
Table of Contents  
Table of Contents ............................................................................................................................................................................ 9 
Glossary of Abbreviations ............................................................................................................................................................. 13 
1.  Background and Rationale ....................................................................................................................................................... 16 
1.1.  Background and Scientific Rationale ................................................................................................................................. 16 
1.2. Rationale for Selection of Investigational Product ............................................................................................................. 16 
1.3. Preclinical Experience ......................................................................................................................................................... 17 
1.4.  Clinical Studies ................................................................................................................................................................... 19 
2.  Study Hypotheses/Objectives .................................................................................................................................................. 21 
2.1.  Hypotheses ........................................................................................................................................................................ 21 
2.2.  Primary Objective .............................................................................................................................................................. 21 
2.3.  Secondary Objectives ......................................................................................................................................................... 21 
2.4.  Exploratory Objectives ....................................................................................................................................................... 22 


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3.  Study Design ............................................................................................................................................................................ 22 
3.1.  Description of Study Design ............................................................................................................................................... 22 
3.2.  Primary Endpoint ............................................................................................................................................................... 24 
3.3. Secondary Endpoints .......................................................................................................................................................... 24 
3.4. Exploratory Endpoints ........................................................................................................................................................ 24 
3.5.  Stratification, Randomization, and Blinding/Masking ....................................................................................................... 25 
3.5.1.  Procedure for Unblinding/Unmasking ....................................................................................................................... 25 
4.  Selection of Participants and Clinical Sites/Laboratories ........................................................................................................ 26 
4.1.  Rationale for Study Population .......................................................................................................................................... 26 
4.2.  Inclusion Criteria ................................................................................................................................................................ 26 
4.3.  Exclusion Criteria ............................................................................................................................................................... 26 
4.4.  Selection of Clinical Sites/Labs ........................................................................................................................................... 27 
5.  Known and Potential Risks and Benefits to Participants ......................................................................................................... 28 
5.1.  Risks of Investigational Product or Intervention as cited in Investigator Brochure .......................................................... 28 
5.2.  Risks of Investigational Product or Intervention as cited in Medical Literature ............................................................... 28 
5.3.  Risks of Other Protocol Specified Medications .................................................................................................................. 28 
5.4.  Risks of Study Procedures .................................................................................................................................................. 28 
5.4.1.  Risks Associated with Stopping the Use of Protocol Prohibited Medications/Therapies.......................................... 28 
5.4.2.  Risks Associated with Skin Swab Collection ............................................................................................................... 28 
5.4.3.  Risks Associated with Physical Exam .......................................................................................................................... 29 
5.4.4.  Risks associated with Questionnaires ........................................................................................................................ 29 
5.4.5.  Risks associated with Blood Collection ...................................................................................................................... 29 
5.5.  Risks Associated with Electronic Data Systems ................................................................................................................. 29 
5.6.  Potential Benefits ............................................................................................................................................................... 29 
6.  Investigational Agent ............................................................................................................................................................... 29 
6.1.  Investigational Agent ......................................................................................................................................................... 29 
6.1.1.  S. hominis A9 Product ................................................................................................................................................ 29 
6.1.2.  Placebo ....................................................................................................................................................................... 31 
6.2.  Investigational Product Accountability .............................................................................................................................. 32 
6.3.  Assessment of Participant Compliance with Investigational Agent .................................................................................. 32 
6.4.  Toxicity Prevention and Management ............................................................................................................................... 32 
6.5.  Premature Discontinuation of Investigational Agent ........................................................................................................ 33 


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7.  Other Medications ................................................................................................................................................................... 33 
7.1.  Concomitant Medications .................................................................................................................................................. 33 
7.1.1  Protocol-mandated ..................................................................................................................................................... 33 
7.1.2  Other permitted concomitant medications ................................................................................................................ 33 
7.2.  Prophylactic Medications .................................................................................................................................................. 33 
7.3.  Prohibited Medications/Therapies .................................................................................................................................... 33 
7.4.  Rescue Medications ........................................................................................................................................................... 34 
8.  Study Procedures ..................................................................................................................................................................... 34 
8.1.  Recruitment Visit ............................................................................................................................................................... 34 
8.2.  Screening Visit (Day -14 to -7)............................................................................................................................................ 35 
8.3.  Pre-treatment Visit (Day -7 ± 1 Day) .................................................................................................................................. 35 
8.4.  Treatment Visit (Day 0) ...................................................................................................................................................... 36 
8.5.  Follow-Up Visits (24 Hour and Days 3, 10, 17 and 24) ....................................................................................................... 37 
8.6. End of Study Phone Visit (Day 31 ± 7 Days) ........................................................................................................................ 37 
8.7. Unscheduled Visits .............................................................................................................................................................. 38 
8.8.  Visit Windows .................................................................................................................................................................... 38 
9.  Mechanistic Assays .................................................................................................................................................................. 38 
9.1.  Skin Swab Collection Procedure ........................................................................................................................................ 38 
9.2.  CFU Quantification of Live Staphylococcus aureus and CoNS ........................................................................................... 39 
9.3.  Quantification of S. hominis A9 CFU .................................................................................................................................. 39 
9.4.  Quantification of Bacterial DNA ......................................................................................................................................... 39 
9.5 Analysis of Antibiotic Sensitivity .......................................................................................................................................... 39 
10.  Biospecimen Storage ............................................................................................................................................................. 40 
11.  Criteria for Participant and Study Completion and Premature Study Termination .............................................................. 40 
11.1.  Participant Completion .................................................................................................................................................... 40 
11.2.  Participant Stopping Rules and Withdrawal Criteria ....................................................................................................... 40 
11.3.  Participant Replacement ................................................................................................................................................. 40 
11.4.  Follow-up after Early Study Withdrawal .......................................................................................................................... 40 
11.5.  Study Stopping Rules ....................................................................................................................................................... 41 
12.  Safety Monitoring and Reporting .......................................................................................................................................... 41 
12.1 Overview ............................................................................................................................................................................ 41 
12.2 Definitions .......................................................................................................................................................................... 41 


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12.2.1 Adverse Event (AE) ..................................................................................................................................................... 41 
12.2.2 Unexpected Adverse Event or Unexpected Suspected Adverse Reaction ................................................................ 42 
12.2.3 Serious Adverse Event (SAE) or Serious Suspected Adverse Reaction ...................................................................... 42 
12.3 Grading and Attribution of Adverse Events ....................................................................................................................... 42 
12.3.1 Grading Criteria .......................................................................................................................................................... 42 
12.3.2 Attribution Definitions ............................................................................................................................................... 43 
12.4 Collection and Recording of Adverse Events ..................................................................................................................... 44 
12.4.1 Collection Period ........................................................................................................................................................ 44 
12.4.2 Collecting Adverse Events .......................................................................................................................................... 44 
12.4.3 Recording Adverse Events ......................................................................................................................................... 44 
12.5 Reporting of Serious Adverse Events and Adverse Events ................................................................................................ 44 
12.5.1 Reporting of Serious Adverse Events to DAIT/NIAID ................................................................................................. 44 
12.5.2 Reporting to Health Authority ................................................................................................................................... 45 
12.5.3 Reporting of Adverse Events to IRB/IEC .................................................................................................................... 46 
12.6 Pregnancy Reporting.......................................................................................................................................................... 46 
12.7 Reporting of Other Safety Information.............................................................................................................................. 47 
12.8 Review of Safety Information ............................................................................................................................................ 47 
12.8.1 Medical Monitor Review ............................................................................................................................................ 47 
12.8.2 DSMB Review ............................................................................................................................................................. 47 
13. Statistical Considerations and Analytical Plan ........................................................................................................................ 48 
13.1. Overview ........................................................................................................................................................................... 48 
13.2.  Endpoints ......................................................................................................................................................................... 48 
13.2.1 Primary Endpoint ....................................................................................................................................................... 48 
13.2.2 Secondary Endpoints ................................................................................................................................................. 48 
13.2.3 Exploratory Endpoints................................................................................................................................................ 48 
13.3.  Measures to Minimize Bias .............................................................................................................................................. 49 
13.4.  Analysis Plan .................................................................................................................................................................... 49 
13.4.1 Analysis Populations .................................................................................................................................................. 49 
13.4.2 Primary Analysis of Primary Endpoint ....................................................................................................................... 49 
13.4.3 Supportive Analyses of the Primary Endpoint ........................................................................................................... 49 
13.4.4 Analyses of Secondary Endpoints .............................................................................................................................. 49 
13.4.5 Analyses of Exploratory Endpoints ............................................................................................................................ 50 
13.4.6 Descriptive Analyses .................................................................................................................................................. 50 


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13.5.  Interim Analyses .............................................................................................................................................................. 50 
13.5.1 Interim Analysis of Primary Endpoint ........................................................................................................................ 50 
13.5.2 Interim Analysis of Safety Data .................................................................................................................................. 50 
13.5.3 Futility Analysis .......................................................................................................................................................... 50 
13.6.  Statistical Hypotheses ...................................................................................................................................................... 50 
13.7.  Sample Size Considerations ............................................................................................................................................. 51 
14.  Identification and Access to Source Data .............................................................................................................................. 51 
14.1.  Source Data ...................................................................................................................................................................... 51 
14.2.  Access to Source Data ...................................................................................................................................................... 51 
15.  Protocol Deviations ................................................................................................................................................................ 51 
15.1.  Protocol Deviation Definitions ......................................................................................................................................... 51 
15.2.  Reporting and Managing Protocol Deviations ................................................................................................................. 51 
16.  Ethical Considerations and Compliance with Good Clinical Practice ..................................................................................... 52 
16.1.  Quality Control and Quality Assurance ............................................................................................................................ 52 
16.2.  Statement of Compliance ................................................................................................................................................ 52 
16.3.  Informed Consent Process ............................................................................................................................................... 53 
16.4.  Privacy and Confidentiality .............................................................................................................................................. 53 
17.  Publication Policy ................................................................................................................................................................... 53 
18.  References ............................................................................................................................................................................. 54 
Appendix A: ADRN Standard Diagnostic Criteria (Version 3.0 09May2014) ................................................................................ 56 
Appendix B: Schedule of Events ................................................................................................................................................... 58 
 
  
Glossary of Abbreviations  
AD  
Atopic Dermatitis  
ADRN  
Atopic Dermatitis Research Network  
AE  
Adverse Event  
AIP  
Autoinducing Peptide  
AMP  
Antimicrobial peptide  
BSA  
Body Surface Area  
CoNS  
Coagulase Negative Staphylococcal Species  


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CFR  
Code of Federal Regulations  
CFU  
Colony-Forming Unit  
CRF  
Case Report Form  
CTCAE  
Common Terminology Criteria for Adverse Events  
DAIT  
Division of Allergy, Immunology, and Transplantation  
DSMB  
Data and Safety Monitoring Board  
eCRF  
Electronic Case Report Form  
FDA  
Food and Drug Administration  
GCP  
Good Clinical Practice  
ICH  
International Conference on Harmonisation  
IGA  
Investigator Global Assessment  
IRB  
Institutional Review Board  
ITT  
Intent To Treat  
L-EASI  
Local Eczema Area and Severity Index  
MCB  
Master Cell Bank  
MOP  
Manual of Procedures  
MRSA  
Methicillin-resistant Staphylococcus aureus  
NBUVB  
Narrow Band Ultraviolet B  
NCI  
National Cancer Institute  
NIAID  
National Institute of Allergy and Infectious Diseases  
NRS  
Numerical Rating Scale  
OVA  
Ovalbumin  
PBS  
Phosphate-Buffered Saline Solution   
PI  
Principal Investigator  
PUVA  
Psoralen Ultraviolet A  
S. aureus  
Staphylococcus aureus  
S. hominis  
Staphylococcus hominis  
SACCC  
Statistical and Clinical Coordinating Center   
 


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SAE  
Serious Adverse Event  
SAR  
Suspected Adverse Reaction  
SCORAD  
Scoring Atopic Dermatitis  
SOP  
Standard Operating Procedure  
SUSAR  
Serious and Unexpected Suspected Adverse Reaction  
TSB  
Tryptic Soy Broth  
USP  
United States Pharmacopeia  
UVA  
Ultraviolet A  
UVB  
Ultraviolet B  
WCB  
Working Cell Bank  


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1. Background and Rationale   
1.1. Background and Scientific Rationale  
Atopic Dermatitis (AD) is a common, chronic, inflammatory skin disease complicated by recurrent bacterial as well as 
viral skin infections (Gao, et al., 2012).  The skin of AD patients is highly susceptible to infections by viruses, bacteria, and 
fungi.  These microorganisms interact with the human immune system to trigger the onset or exacerbate the disease.  
Studies suggest that 7-10% of AD patients have difficulty containing infections caused by viruses, including herpes 
simplex virus, vaccinia virus (causing eczema vaccinatum), and molluscum contagiosum virus (Howell, et al., 2006).  
Preliminary data in the Atopic Dermatitis Research Network (ADRN) Registry study indicate 40% of AD subjects, and only 
2% of non-atopics are Staphylococcus aureus (S. aureus) colonized on lesional or non-lesional skin.  These observations 
are consistent with the understanding that patients with AD have defects in innate and adaptive immune responses and 
thus have a general defect in the control of skin infections.  
  
Several lines of experimental evidence support the concept that the multiple defects in epithelial barrier function in AD 
promote dysbiosis, a state of microbial imbalance, and that this dysbiosis then promotes the immunological disorder 
characteristic of AD.  Dysbiosis in AD has been shown to be closely associated with disease severity (Leyden, et al., 
1974).  Antibiotic therapy is a mainstay for treatment of bacterial infections; however, antibiotic treatment 
nonspecifically kills bacteria and may induce additional dysbiosis.  In addition, long-term antibiotic treatments generate 
resistant bacteria.  Human antimicrobial peptides (AMPs) such as defensins and cathelicidins are important mediators to 
maintain the balance of microflora on the skin.  Our studies indicate that the innate immune defense of the skin is 
mediated not only by AMPs produced by an individual’s own cells, but also by antimicrobial molecules contributed by 
commensal coagulase negative staphylococcus species (CoNS), such as S. epidermidis and S. hominis (Nakatsuji, et al., 
2017; Dorschner, et al., 2001; Murakami, et al., 2002; DiNardo, et al., 2003; Zaiou, et al., 2003).  More recently, we have 
found that autoinducing peptide (AIP) produced by skin commensal CoNS suppresses production of virulence factors, 
including Phenol-soluble modulin-alpha, in S. aureus (Williams, et al., 2019).  Our clinical research also suggests that AD 
skin colonized by S. aureus has a lower frequency of these antimicrobial Staph strains (CoNS AM+) and CoNS strains with 
capacity to produce AIP compared to normal skin (Nakatsuji, et al., 2017).  Studies have also shown that AD patients 
colonized by S. aureus have more severe disease states and are prone to more frequent flares of their AD (Leyden, et al., 
1974; Kong, et al., 2012).  We hypothesize that the skin of patients with AD selectively inhibits the survival of beneficial 
bacteria (CoNS AM+) that inhibit S. aureus, and that an increase in the amount of CoNS AM+ to the level found on most 
healthy skin would normalize the balance of microflora in AD patients, thereby improving abnormal immune reactions 
by AD skin.  The goal of this study is to understand why beneficial bacteria are lost in AD, and to better understand the 
factors involved in survival of beneficial bacterial strains (CoNS AM+).   
1.2. Rationale for Selection of Investigational Product   
The functions described in Section 1.1 for the microbiome have focused on S. epidermidis and S. aureus bacterial 
species.  However, we have made a highly innovative discovery of newly recognized species such as S. hominis, as well as 
newly identified strains of S. epidermidis from the human skin microbiome that produce previously unknown 
antimicrobials that will selectively kill pathogens such as S. aureus while not harming the resident commensals.  Because 
of this, these commensal bacteria can provide selective antimicrobial protection against pathogens on the skin surface 
yet maintain diversity.  
One example of such protective bacteria is a strain of S. epidermidis that produces Phenol Soluble Modulins gamma and 
delta (Cogen, et al., 2010a; Cogen, et al., 2010b).  We have shown that these helical peptides are potent against S. 
aureus in vitro and on human skin.  More recently, with the prior support of the National Institute of Allergy and 






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Figure 1.3c. Deletion of lantibiotic from a CoNS-AM+ S. hominis strain converts it to CoNS-AM- and improves its 
resistance to human AMP  
  
(A) lantibiotics-alpha-hogocidin and beta-hogocidin genes were deleted by allelic reaction with pKOR1 plasmid in colony 2 (arrow head).  (B) Colony 2 did not produce antimicrobial activity 
against S. aureus.  Antimicrobial activity is visualized by zone of inhibition around (arrow) bacteria colony (white dot). (C) Shows that expression of the lantibiotics makes S. hominis A9 more 
susceptible to killing by LL-37.  The alpha and beta lantibiotic mutant, or the parent WT S. hominis A9, was cultured in the presence of indicated concentrations of the human AMP LL-37 at 
30oC for 24 hours.  Relative growth was calculated from OD600.(Unpublished data, Gallo lab-UCSD)  
  
  
The skin pathogen Staphylococcus aureus has been reported to have highly efficient mechanisms for horizontal gene 
transfer between strains (Chen, et al., 2018; Humphrey, et al., 2021). Similar mechanisms of transfer have not been 
reported for Staphylococcus hominis. Additionally, in our analysis of ShA9, there has been no evidence of S. aureus 
lateral gene transduction elements in ShA9 and no evidence ShA9 has acquired virulence or antibiotic resistance genes 
via conjugative transposon from other species.  Plasmids for antibiotic resistance genes or virulence factors also have 
not been found in our analysis of S. hominis. However, we did identify the presence of two prophages containing a 
betalactamase homolog in the ShA9 DNA sequence (unpublished data, Gallo lab-UCSD).  The antibiotic resistance profile 
present in ShA9 to beta-lactam antibiotics, tetracycline, and erythromycin is also already ubiquitously present in the 
healthy human skin microbiome (Oh, et al., 2014; Oh, et al., 2016; and Baron, et al., 2018); therefore, the effect of this 
occurrence is unclear. Due to the theoretical possibility of transfer of antibiotic resistance from ShA9 to Staphylococci 
present on the skin of study participants, we will investigate the antibiotic sensitivity to penicillin G, tetracycline, and 
erythromycin in CoNS isolated from skin swabs at baseline and at the last swab collection from lesional skin.  
  
1.4. Clinical Studies  
In a previous study conducted under this IND (Protocol ADRN-08), we were able to establish that dermal application of 
ShA9 was safe to use on humans.  ADRN-08 was a phase 1 study of 54 S. aureus positive AD subjects randomized 2:1 
active to vehicle (29 female and 25 male, ages 18-57; ClinicalTrials.gov, NCT03151148).  We found application of S. 
hominis A9 to be safe in comparison to vehicle, with no difference in the per participant adverse event (AE) rate 
between active drug and vehicle (p=0.075).  In addition, there was a statistically significant decrease in live S. aureus 
(p<0.001), on participants treated with ShA9 compared to vehicle (Nakatsuji, et al., 2021).  
In our previous Phase 1 trial, we obtained data that led us to test the hypothesis that the elimination of ShA9 from 
lesional skin of AD participants was due to susceptibility of this strain to killing by antimicrobials produced by inflamed 
skin (Nakatsuji, et al., 2021). We further hypothesized that this susceptibility is a consequence of the expression of the 
lantibiotics produced by ShA9 but not by other strains of S. hominis that do not produce lantibiotics. This hypothesis was 
supported by the pre-clinical observations shown in Figure 1.3 a-c that CoNS-AM+ strains survive less well on inflamed 


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mouse skin, are more susceptible to killing by the human AMPs, such as cathelicidin and β -defensins -2 and-3 present 
on inflamed skin of patients with AD, and ShA9 increases survival upon deletion of its lantibiotics. To further test this 
hypothesis on human skin in the clinical setting, we measured abundance of ShA9 on the skin of participants from the 
Phase 1 trial under Protocol ADRN-08.   We measured abundance of ShA9 on inflamed and non-inflamed skin. ShA9 
abundance was measured by multiple approaches, including direct counting of total CoNS CFUs of live bacteria grown on 
selective agar plates, abundance of ShA9 DNA by qPCR, and measurement of mRNA by RT-qPCR for candidate ShA9 gene 
expression.  Total CoNS CFUs were used as a surrogate measurement of live ShA9 CFU, since ShA9 is not specifically 
measurable by colony counting methods. However, since ShA9 is a type of CoNS, and the abundance of ShA9 applied 
greatly exceeded other CoNS recoverable from AD skin, this estimation was helpful to define live compared to dead 
bacteria. Furthermore, measurements by qPCR of ShA9 DNA and mRNA provide a more specific method to validate this 
assumption.  On Day 0, an initial swab was collected one hour after application of ShA9 or vehicle.   Both lesional and 
non-lesional skin showed a 3- to 4-log increase in live CoNS on the skin one hour after the first application of ShA9 but 
no change after application of the vehicle alone (Figure 1.4a).  However, a difference in CoNS survival was observed 
between lesional and non-lesional sites at Days 4 and 7.  At these time points, skin swabs were collected approximately 
four hours after the morning application.  Swabs from lesional skin collected at these later times on Days 4 and 7 
showed that live CoNS had decreased to a level similar to baseline, and no significant difference was detected in 
comparison to that from vehicle-treated subjects.   This suggested the longer period between the morning application of  
ShA9 and the skin sampling resulted in a decrease in the live ShA9 recoverable by swab from lesional skin.  In contrast, 
CoNS recovered from non-lesional skin was greater on both Day 4 and Day 7 on participants treated with ShA9 
compared to vehicle (Figure 1.4a).  These data support our preclinical observations that CoNS-AM+ bacteria survive less 
well in an inflamed skin environment that will have expression of human AMPs.   
Figure 1.4a.  CoNS-AM+ persist on non-lesional skin but not on inflamed skin of AD adult participants.   
  
  
  
Change in live coagulase-negative Staphylococci (CoNS) recovered by skin swab from patient’s skin compared to baseline at indicated time points after application of ShA9 or vehicle.  ShA9  
(n=35), vehicle (n=17).  Data represent Mean ± SEM.  * p< 0.05, ** p< 0.01, *** p< 0.001, by t-test.   (Nakatsuji, et al., 2021)  
  
To determine if other forms of skin inflammation also alter the CoNS-AM+ population, we have also recently measured 
CoNS-AM+ on patients with psoriasis.  We reasoned that since psoriasis patients have even higher amounts of AMPs on 
their skin than AD subjects, the host innate immune system of psoriasis would select against CoNS-AM+.  This would 
occur despite the contrast in T-cell repertoire between AD and psoriasis (Guttman-Yassky, et al., 2011a; Guttman-Yassky, 
et al., 2011b).  Our preliminary data suggest patients with psoriasis also have a deficiency of CoNS-AM+ (Figure 1.4b).  
Thus, although AD and psoriasis are driven by distinct immune responses, the loss of CoNS-AM+ strains was observed on 
the skin of both diseases.  These data strongly support our hypothesis that AMPs select against CoNS-AM+.  We further 
hypothesize that since psoriasis has high mammalian AMPs, this level is sufficient to resist dysbiosis driven by S. aureus.  




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2.4. Exploratory Objectives  
1. To assess the association of S. hominis A9 estimated CFU on the skin of the ventral arm contralateral to the arm 
of application and face  
2. To assess the duration of survival of CoNS as measured by CFU during a maximum of 24 days after the 
application of S. hominis A9 on lesional ventral arm skin of AD participants negative for S. aureus (AD SA-)  
3. To assess the duration of survival of CoNS as measured by CFU during a maximum of 24 days after the 
application of S. hominis A9 on non-lesional ventral arm skin of AD participants negative for S. aureus (AD SA-)  
4. To compare the duration of survival of CoNS as measured by CFU on lesional skin between AD SA+ vs AD SA- 
participants  
5. To compare the duration of survival of CoNS as measured by CFU on non-lesional skin between AD SA+ vs AD 
SA- participants  
6. To compare the duration of survival of CoNS as measured by CFU between lesional and non-lesional skin on AD 
SA+ participants  
7. To compare the duration of survival of CoNS as measured by CFU between lesional and non-lesional skin on AD 
SA- participants  
8. To compare disease severity measures (local Eczema Area and Severity Index [L-EASI], SCORing Atopic Dermatitis  
[SCORAD] and local Pruritus Numerical Rating Scale [NRS]) between Day 0 and the last in-clinic visit among AD 
SA+ participants, and independently, among AD SA-  
9. To determine the antibiotic sensitivity of the skin CoNS microbiome to penicillin G, tetracycline, and 
erythromycin before and after treatment with ShA9  
3.  Study Design  
3.1. Description of Study Design   
Protocol ADRN-13 is a Phase 1 pilot, open label, single center trial designed to assess the kinetics of S. hominis A9 
survival in adults with moderate-to-severe atopic dermatitis on the ventral arms who are culture positive or negative for 
Staphylococcus aureus colonization.  Total CoNS CFU will be used to assess S. hominis A9 survival since this specific 
species cannot be distinguished by culture and it is expected to be the predominant CoNS species after the treatment 
application.   
A minimum of 20 AD subjects (13 S. aureus positive and 7 S. aureus negative), 18 to 80 years of age, will have a single 
application of S. hominis A9 applied to their right or left ventral arm (wrist to upper arm); a single application of placebo 
will be applied to the contralateral arm.  If any participants withdraw or are withdrawn prior to achieving CoNS 
elimination (CoNS CFU below baseline density measured before application of ShA9 + 100 CFU/cm2 ), additional 
participants may be enrolled as necessary to obtain a modified intent to treat (ITT) sample as defined in Section 13.4.1.  
Potential participants will be recruited in person or over the phone and assessed for eligibility.  Potentially eligible 
participants, who have no obvious characteristics making them ineligible and who are interested in study participation, 
will be invited to complete an in-clinic Screening Visit.  During the Screening Visit, participants will provide informed 
consent for study participation.  Consented participants will then be further assessed for full study eligibility through the 
collection of medical history, including medication/therapy use, a physical exam, and assessment of AD severity.  
Participants who meet all eligibility requirements, including medication and therapy washouts (no topical AD treatments 


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or prescription moisturizers on the ventral arms or face, and no oral or topical antibiotics, or bleach baths in the last 7 
days, and no oral steroid therapy, phototherapy, or regular tanning bed use [more than 2 visits per week] in the last 28 
days), may complete the Pre-treatment Visit on the same day of the Screening Visit.  Participants who require a 
medication/therapy washout of more than 14 days will be required to rescreen.   
Screened and eligible participants will complete a Pre-treatment Visit (Day -7 ± 1 Day), approximately 7 days prior to their 
Day 0 Visit.  During the visit, target lesional and non-lesional sites, each measuring at least 21 cm2 total, will be identified 
on the participant’s right and left ventral arms.  The sites will be photographed for reference at the participant’s future 
visits, and swabs for S. aureus screening by culture, S. aureus quantification by PCR, CoNS CFU, and S. hominis A9 
estimated CFU will be collected from the identified sites.  Participants will also be provided Cetaphil lotion and Dove soap 
during this visit, for use during the week prior to their Treatment Visit (Day 0).   
Participants who complete the pre-treatment phase (7 ± 1 days total) and meet enrollment criteria for AD SA+ or AD SA-, 
will come back to clinic for their Treatment Visit (Day 0).  Participants must be cultured to determine S. aureus 
colonization status.  Given the rates of S. aureus colonization, it is likely that enrollment will be complete for the AD SA- 
group before the AD SA+ group.  Once the enrollment goals for AD SA+ or AD SA- are met, additional participants will be 
identified as screen failures. During the Treatment Visit, the participant will have ShA9 applied to their right or left 
ventral arm and placebo applied to their contralateral ventral arm.  The assignment of ShA9 and placebo to the 
dominant and non-dominant arms will be randomized.  Prior to the application of the ShA9 and placebo treatments, skin 
swabs will be collected from the lesional and non-lesional sites on the ventral arms and one non-lesional site on the 
participant’s face.  A blinded clinical team member will apply the designated treatment to each arm, and additional 
swabs will be collected from the arms 15 minutes and 1, 2, 4, and 6 hours after application.  Participants will return to 
the clinic 24 hours after application (24 Hour Visit) and on Days 3, 10, 17, and 24 for the assessment of AEs and the 
collection of skin swabs from the identified target sites, as needed.  After completion of the Day 3 Visit, a participant will 
not be required to complete the remaining follow up clinic visits (Day 10 through Day 24) if their lesional swabs from 
both arms (right and left) are CoNS negative, defined as CoNS below baseline density measured before application of 
ShA9 + 100 CFU/cm2.  All randomized participants will complete a final End of Study Phone Visit on Day 31 to assess 
safety and disease status.   
A minimum of 13 S. aureus positive (defined as S. aureus CFU ≥ 100 CFU/cm2 for lesional swabs from both arms) and a 
minimum of 7 S. aureus negative (defined as S. aureus CFU < 100 CFU/cm2 for lesional swabs from both arms)  
participants with moderate-to-severe AD will be enrolled to receive study treatment in this trial.  Based on preliminary 
data from the ADRN Registry study (unpublished data), we estimate that approximately 40% of AD participants will be S. 
aureus positive on their lesional skin, and we may need to screen (including pretreatment) approximately 33 participants 
to enroll 13 AD S. aureus positive participants, as colonization is determined once skin swabs are collected at the 
Pretreatment Visit.  We anticipate it will take approximately 18 months to reach the recruitment and enrollment goals 
and to complete participant follow up for this trial.  The study flow diagram is provided in Figure 3.1.    
Figure 3.1 Study Design Schematic  




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2. The duration of survival, measured as the time needed for CoNS CFU to drop below baseline density measured 
before application of ShA9 + 100 CFU/cm2 on the lesional ventral arm skin of AD participants negative for S. 
aureus (AD SA-)  
3. The duration of survival, measured as the time needed for CoNS CFU to drop below baseline density measured 
before application of ShA9 + 100 CFU/cm2 on the non-lesional ventral arm skin of AD participants negative for S. 
aureus (AD SA-)  
4. Comparison of time needed for CoNS CFU to drop below baseline density measured before application of ShA9 +  
100 CFU/cm2 on the lesional and non-lesional, separately, ventral arm skin between AD participants positive (AD 
SA+) vs negative (AD SA-) for S. aureus   
5. Comparison of time needed for CoNS CFU to drop  below baseline density measured before application of ShA9 
+ 100 CFU/cm2 on lesional vs non-lesional skin of AD participants positive (AD SA+) and negative (AD SA-) for S. 
aureus, separately  
6. Comparison of the L-EASI score of the ventral arms at Day 0 vs the last in-clinic visit among AD SA+, and 
independently, among AD SA-    
7. Comparison of the SCORAD score at Day 0 vs the last in-clinic visit among AD SA+, and independently, among AD  
SA-    
8. Comparison of the local Pruritus NRS of the ventral arms at Day 0 vs the last in-clinic visit among AD SA+, and 
independently, among AD SA-   
9. Comparison of the proportion of antibiotic-sensitive CoNS to penicillin G, tetracycline, and erythromycin isolated 
from skin swabs at baseline and after ShA9 treatment, at the last swab collection from lesional skin  
3.5. Stratification, Randomization, and Blinding/Masking  
Arm dominance may affect the spread of ShA9 to the contralateral arm; therefore, the design will stratify by arm 
dominance, with the dominant arm being the right arm for right-handed participants, and the left arm for left-handed 
participants.  A minimum of 13 participants with S. aureus colonized lesions on their ventral arms and a minimum of 7 
participants with S. aureus negative lesions on their ventral arms will be randomized to one of two treatment groups: 1) 
ShA9 dominant and placebo non-dominant or 2) ShA9 non-dominant and placebo dominant.  It is anticipated that 
enrollment will be higher for right-handed participants as that is the more dominant arm in the general population; 
enrollment will not target equal numbers of right-handed and left-handed participants.  Randomization will be 
performed centrally at the Statistical and Clinical Coordinating Center (SACCC) using a stratified randomization design.   
This is an open-label study, as all participants will receive active and placebo treatment.  However, investigators, all 
investigational site staff, including those responsible for preparing/administering the investigational product, and all 
participants in this study will be blinded as to which arm received which treatment.  Laboratory staff performing the 
mechanistic assays will be unblinded to which arm received which treatment, to ensure the correct dilutions can be 
applied to each sample prior to completing assays for quantification of live bacteria by culture (S. aureus and CoNS) and 
bacterial DNA (S. hominis A9) by qPCR (Refer to Section 9).  
3.5.1. Procedure for Unblinding/Unmasking  
If a clinically significant event occurs and knowledge of the treatment assignment for a given arm is required, the 
study treatment may be unblinded.  Unblinding must be approved by the study Medical Monitor unless an 
immediate life-threatening condition has developed and the Medical Monitor is not accessible.  The site 
investigator will notify the Medical Monitor of the unblinding event within 1 business day, following the 
emergency unblinding.  The emergency unblinding will also be reported to the Data and Safety Monitoring 
Board (DSMB).  


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A full account of the event will be recorded, including the date and time of the unblinding, the reason for the 
decision to unblind, the name of the individual who made the decision, and the names of the Medical Monitor 
and others who were notified.  The reasons for unblinding of a participant’s treatment will be included in the 
final study report.  
Unblinding will also occur for Investigational New Drug Safety Reports that will be reported to the Food and 
Drug Administration (FDA), DSMB, and Institutional Review Board (IRB) as specified in the current FDA IND 
Safety Reporting Guidance.   
Unblinding the study due to an approved interim analysis, final analysis, or study termination will require 
written approval from NIAID.  
4. Selection of Participants and Clinical Sites/Laboratories  
4.1. Rationale for Study Population  
In atopic dermatitis, a state of microbial imbalance known as dysbiosis occurs and is closely associated with disease 
severity.  An important characteristic of the dysbiosis in AD is an increased abundance of S. aureus and a decrease in 
overall bacterial diversity.  Several lines of experimental evidence support the concept that the multiple defects in 
epithelial barrier function in AD promote dysbiosis and that this dysbiosis then promotes the immunological disorder 
characteristic of AD (Paller, et al., 2019).  Previous results have further suggested that S. hominis A9 produces 
bacterocins that kill S. aureus (Nakatsuji, et al., 2017).  These characteristics, dysbiosis, and immunological disorder 
make AD patients most suitable for the transplant of beneficial skin microbiome species, such as S. hominis A9, to help 
regulate the imbalance.  In this study, we will examine the survival of S. hominis A9 transplanted bacteria on lesional and 
non-lesional skin of AD subjects with and without S. aureus colonization.  The inclusion of AD subjects without S. aureus 
colonization will permit assessment of the potential of S. aureus to produce factors that limit the survival of S. hominis 
A9.  Each participant will receive S. hominis A9 on one arm and placebo on the contralateral arm which will serve as a 
control.  This study will be limited to adult AD patients, ages 18-80 years.  
4.2. Inclusion Criteria  
Individuals who meet all of the following criteria are eligible for enrollment as study participants:  
1. Participant must be able to understand and provide informed consent.  
2. Male or female participants 18 to 80 years of age, inclusive at time of the Screening Visit.   
3. Meet ADRN Standard Diagnostic Criteria (Appendix A) for active AD.  
4. At least 21 cm2 of lesional and 21 cm2 of non-lesional skin on both the right and left ventral arms.  The required 
area (lesional or non-lesional) may be one contiguous area or may encompass multiple areas with a total 
cumulative area of 21 cm2.  
5. An Investigator Global Assessment (IGA) score, on the ventral arms of at least moderate severity.  
6. Body surface area (BSA), as measured by Mostellar BSA Calculator, between 1.26 m2 and 2.25 m2.  
7. If female of child bearing potential, must agree to remain abstinent (refrain from heterosexual intercourse) or 
use acceptable contraception (e.g.  oral contraceptives, intrauterine device [IUD], barrier method with 
spermicide, surgically sterilized partner, Depo-Provera, Norplant, NuvaRing, or hormonal implants) for the 
duration of study participation.  
4.3. Exclusion Criteria  
Individuals who meet any of these criteria are not eligible for enrollment as study participants:  


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1. Inability or unwillingness of a participant to give written informed consent or comply with study protocol  
2. Pregnant or lactating females.   
3. Active bacterial, viral, or fungal skin infections.  
4. Any noticeable breaks or cracks in the skin on the target areas of investigational product application, including 
severely excoriated skin or skin with open or weeping wounds suggestive of an active infection or increased 
susceptibility to infection.  
5. Sensitivity to or difficulty tolerating Dove fragrance-free bar soap, Cetaphil® lotion, alcohol-based cleaners, 
glycerol, or soy products.  
6. Participants with Netherton’s syndrome or other genodermatoses that result in a defective epidermal barrier  
7. Any participant who is immunocompromised (e.g. history of lymphoma, Human Immunodeficiency Virus 
[HIV]/Acquired Immunodeficiency Syndrome [AIDS], Wiskott-Aldrich Syndrome), has an immune system 
disorder (e.g. autoimmune disease), or is using a systemic immunosuppressant (e.g. systemic corticosteroids, 
cyclosporine, methotrexate).   
8. Any participant with current malignant disease (with the exception of non-melanoma skin cancer in an area not 
affected by treatment).  
9. Participants with a history of psychiatric disease or history of alcohol or drug abuse that would interfere with the 
ability to comply with the study protocol.  
10. Past or current medical problems or findings from physical examination or laboratory testing that are not listed 
above, which, in the opinion of the investigator, may pose additional risks from participation in the study, may 
interfere with the participant’s ability to comply with study requirements or that may impact the quality or 
interpretation of the data obtained from the study.  
11. Ongoing participation in another investigational trial or use of investigational drugs within 8 weeks, or 5 halflives 
(if known), whichever is longer, of the Screening Visit.  
12. Treatment with non-steroid systemic immunosuppressant within 6 months of the Screening Visit.  
13. Treatment with Dupilumab within 16 weeks of the Screening Visit.  
14. Treatment with oral or injectable therapy for AD (excluding oral steroids) within 5 half-lives (if known) or 16 
weeks before the Screening Visit, whichever is longer.   
15. Participants with close contacts (e.g. spouse, children, or members in the same household) that have severe 
barrier defects or are immunocompromised.  
16. Use of topical (including steroids and calcineurin inhibitors) AD treatments on the ventral arms or face within 7 
days of the Treatment Visit; Use of topical steroids on areas outside of where investigational product is to be 
applied or swabbing is to be performed may be permitted, per investigator discretion.  
17. Treatment with prescription moisturizers classified as medical device (e.g., Atopiclair®, MimyX®, Epiceram®, etc.) 
on the ventral arms or face within 7 days of the Treatment Visit; Use on areas outside of where investigational 
product is to be applied or swabbing is to be performed is permitted.  
18. Use of any oral or topical antibiotic within 7 days of the Treatment Visit.  
19. Participants who have taken a bleach bath within 7 days of the Treatment Visit.  
20. Use of any oral steroid therapies within 28 days of the Treatment Visit.  
21. Any phototherapy for skin disease (such as narrow band ultraviolet B [NBUVB], ultraviolet B [UVB], ultraviolet A1 
[UVA1], psoralen + UVA [PUVA]) or regular use (more than 2 visits per week) of a tanning bed within 28 days of 
the Treatment Visit.  
4.4. Selection of Clinical Sites/Labs   
This study is being conducted at the University of California, San Diego (UCSD), which was a clinical site for the previous  
Phase I Targeted Microbiome Transplant in Atopic Dermatitis study, ADRN-08, conducted under this IND  


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(ClinicalTrials.gov, NCT03151148).   In addition, UCSD is the manufacturer of the S. hominis A9 being used for this study 
and will perform assays for quantification of live bacteria by culture (S. aureus and CoNS) and bacterial DNA (S. hominis 
A9) by qPCR (Refer to Section 9).  UCSD has optimized these assays and has successfully deployed them for previous 
studies.   
5. Known and Potential Risks and Benefits to Participants  
5.1. Risks of Investigational Product or Intervention as cited in Investigator Brochure  
There is a theoretical risk of skin infection associated with the application of S. hominis A9, although the likelihood of 
participants developing a skin infection is low given the applied product will only include nonpathogenic bacteria that 
have AMP-producing abilities.  Furthermore, we will be applying these bacteria topically and will examine all participants 
prior to application to ensure that they do not have any cracks or excoriated skin on their arms.  Theoretical risk of 
contact dermatitis to glycerol is also possible, both with the S. hominis A9 and the placebo.  We will avoid application of 
the product to the hand since the hand would be a likely source of spreading these bacteria onto other surfaces that 
other people may contact.  Participants will be excluded from participating if they have close contacts (e.g. spouse, 
children, or members in the same household) that have severe skin barrier defects or are immunocompromised, per 
study exclusion criteria.   
Should an infection occur, the participant would be treated according to standard treatment, including antibiotics (for 
infection) and ointment (for irritation and dryness).  Contact allergic reactions to glycerol will be treated with 
midpotency topical steroids, such as fluocinonide ointment 0.05%.  Refer to Section 7.4 for more information on rescue 
medications.  
5.2. Risks of Investigational Product or Intervention as cited in Medical Literature  
Not applicable  
5.3. Risks of Other Protocol Specified Medications  
Theoretical risk of contact dermatitis to Cetaphil is possible and will be treated with mid-potency topical steroids, such 
as fluocinonide ointment 0.05%.  Refer to Section 7.4 for more information on rescue medications.  
In the event of infection, the elected rescue treatment will follow guidelines specified in Section 7.4.  There is theoretical 
risk associated with the use of topical or oral antibiotics as a rescue medication.  Specific risks will vary between 
treatments.  In general, risks of antibiotics are hypersensitivity, urticaria, nausea, vomiting, diarrhea, sun sensitivity, 
anaphylaxis, and death.  
5.4. Risks of Study Procedures   
5.4.1. Risks Associated with Stopping the Use of Protocol Prohibited Medications/Therapies  
AEs associated with stopping the use of protocol-prohibited medications/therapies may include worsening of 
the condition being treated and will be reported as such.  In an effort to minimize these risks, participants with 
severe AD who may have difficulty tolerating periods without medication/therapy use will be excluded from 
participating, per study exclusion criteria.  
5.4.2. Risks Associated with Skin Swab Collection  
There are no significant risks associated with skin swab collection.  


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5.4.3. Risks Associated with Physical Exam  
There are no known risks associated with the physical exam.  
5.4.4. Risks associated with Questionnaires  
There is a possibility that participants may find questions too personal.  Participants may refuse to answer any 
questions that make them feel uncomfortable.  There is also a possibility that a participant’s answers may be 
read by others; however, participants’ records are carefully protected so this is very unlikely.  See Section 16.4 
for more information on confidentiality.  
5.4.5. Risks associated with Blood Collection  
There is no scheduled blood draw, but blood may be collected during an Unscheduled Visit per investigator 
discretion.  Risks associated with drawing blood include possible pain when the needle is inserted, as well as 
bleeding, bruising and/or infection at the puncture site.  Some people may experience lightheadedness, nausea, 
or fainting.  There is a potential for slight psychological stress from the procedure.  If psychological stress is too 
much in the opinion of the participant or the physician/nurse, the procedures will be halted.  NIH guidelines for 
blood collection (amount and frequency based on age) will be followed.  
5.5. Risks Associated with Electronic Data Systems  
It is possible that computer/data management systems could be maliciously attacked and personal information as well 
as other study documents compromised.  There is also the potential of accidental loss of privacy during the consenting 
and enrollment process.  To minimize or prevent this from happening, standard procedures such as using coded forms, 
locked files, and encryption at rest and in transit for all data will be employed.  Investigators and the appropriate study 
team members will complete training courses in secure data management/privacy.  Finally, the Rho SACCC data systems 
are fully compliant with Federal regulations and have up-to-date virus and malware protection.  
5.6. Potential Benefits  
There may or may not be any direct benefits for the participants who elect to enroll in this study.  One potential benefit 
is that the investigational product may improve the participant’s AD; however, there is no guarantee that the 
investigational product will help the participant’s condition.  The participant’s skin condition may even get worse by 
withholding his/her previous/regular AD treatment.   
The results of this study may also provide identification and/or validation of new targets for the future development of 
therapeutics for participants with AD, as well as increase current knowledge on the ability to transform the cutaneous 
microbiome, which could lead to potential therapeutic strategies for treating a variety of inflammatory skin conditions.  
6. Investigational Agent  
6.1. Investigational Agent  
6.1.1. S. hominis A9 Product  
 
6.1.1.1.  
Formulation, Packaging, and Labeling   
The S. hominis A9 product will be manufactured and packaged at UCSD.  The UCSD Manufacturing Lab 
has isolated Staphylococcus hominis strain A9 from the skin surface of a healthy donor.  This S. hominis 
A9 strain produces potent antimicrobial activity against S. aureus.  UCSD has characterized antimicrobial 
peptides produced by this strain with protein purification, protein sequencing, and genomic sequencing 
approaches.  The UCSD Manufacturing Lab has established the Master Cell Bank (MCB) of the S. hominis 


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A9 strain, and it is stored in a locked -80°C freezer at UCSD.  The Working Cell Bank (WCB) has been 
prepared from a single colony of the MCB and stored in a locked -80°C freezer at UCSD.  
Each batch of the S. hominis A9 product will be prepared from a single-use vial from the WCB.  A vial of 
the S. hominis A9 from the WCB will be thawed and cultured in United States Pharmacopeia (USP) grade 
animal-free tryptic soy broth (TSB) media at 37°C to expand the number of S. hominis A9.  Bacterial 
concentration will be assessed, after culturing, by OD600. One unit of OD600 is comparable to 5x107 
CFU/ml. The cultured bacteria will then be washed twice with 100 mL of USP grade normal saline to 
remove the animal-free TSB.  Based on pre-clinical data, proteins are undetectable in the supernatant 
after the second wash.  Bacterial density of the resuspension will be measured by OD600.  
Washed bacteria will be mixed under sterile conditions in 85% phosphate-buffered saline solution (PBS) 
containing 15% glycerol to achieve a density similar to the concentration of microbes on normal skin of 
1 x 105 CFU/cm2.   Each batch of formulated product will be assigned a unique lot number and will 
undergo a visual test for appearance, potency testing by bacterial concentration testing and radial 
diffusion testing, and identity testing by PCR for the hogocidin gene.  Aliquots of final S. hominis A9 
product will be transferred into a sterile dispenser with a spray pump (10 mL/dispenser) and sealed in a 
sterile laminar-flow hood.  The dispensers will be grouped in boxes and labeled with the lot number and 
box ID.  Packaged boxes will be stored in a -80°C freezer at UCSD until transfer to the clinical site.  After 
packaging, 2 dispensers from each lot will be randomly selected, and potential pathogens will be 
screened for Microbial Limits Testing in accordance with USP <61> and <62> testing, visual appearance 
testing, potency testing by bacterial concentration testing and radial diffusion testing, and identity 
testing by PCR for the hogocidin gene.  The final S. hominis A9 product will be formulated at a S. hominis 
A9 concentration range from 1 x 108 CFU of bacteria/mL to 4 x 109 CFU of bacteria/mL in PBS-Glycerol.  
Potency testing will be performed by both the radial diffusion assay and bacterial concentration testing.  
Potency of S. hominis A9 product is acceptable if 4 out of 5 S. hominis A9 colonies grown from the 
product show S. aureus inhibition zones greater than 1 mm in diameter.  Bacterial concentration of the 
product will be checked by plating a known quantity of the S. hominis A9 product on an agar plate and 
incubating the plate overnight at 37°C.  The number of viable colonies will be counted the following day.  
All colonies will be checked to ensure they show the same morphology.  Only S. hominis A9 that has a 
final concentration of bacteria that ranges from 1 x 108 CFU of bacteria/mL to 4 x 109 CFU of bacteria/mL 
in PBS-Glycerol will be used in the clinical trial.  
Identity testing will be confirmed by the presence of the hogocidin-α gene by PCR.   
 
6.1.1.2.  
Dosage, Preparation, and Administration  
6.1.1.2.1. Dosage - Targeted Density of Transplanted CoNS  
There is considerable variability in the abundance of CoNS bacteria on human skin.  A 
recent unpublished study by the UCSD laboratory has found CoNS bacteria with 
antimicrobial activity can be detected at a range of abundance between 1 x 102 CFU/cm2 
and 1 x 107 CFU/cm2.  Therefore, the application of CoNS to a final density between 1 x 
102 CFU/cm2 and 1 x 107 CFU/cm2 is the target goal that will be within the density 
typically observed on healthy human skin and can be considered safe.  This target 
density for topically applied CoNS bacteria has also been found safe and effective at 


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inhibiting S. aureus  in prior studies of both autologous microbiome transplant 
(Nakatsuji, et al.,2017) and after topical application of S. hominis A9 (Nakatsuji, et al., 
2021).  
Using the body surface area calculations of Mosteller (Mosteller, 1987), we estimate 
that the smallest potential participant of 4’10” and 85 pounds has a BSA of 1.26 m2.  The 
largest participant of 6’3” and 210 pounds has a BSA of 2.25 m2.  Using the Rule of 
Nines, each arm is estimated as 9% of the total BSA.  Therefore, the ventral surface 
would be 4.5% of the total BSA.  Since one hand is 1% of BSA, the ventral surface of one 
arm not including the hand would be 3.5% of BSA.  Thus, the smallest participant has an 
area on their ventral arm of 441 cm2, and the largest participant 787 cm2.  
The investigational product will be formulated at a final concentration of bacteria that 
ranges from 1 x 108 CFU of bacteria/mL to 4 x 109 CFU of bacteria/mL, and a total 
volume of approximately 1 mL of product will be applied to each participant’s arm once.  
To dispense approximately 1 mL, 6 pumps will be applied per dose (160 µL/pump).  The 
acceptable dosage range will be between 2 x 103 CFU/cm2 and 1 x 107 CFU/cm2 to 
account for variability in participant surface area, bacteria survival, and loss of product 
during application.  The calculated concentration for the arms will be 1.27 x 105 CFU/cm2 
and 9.07 x 106 CFU/cm2 for BSAs of 1.26 m2 and 2.25 m2, respectively.  This range is 
within the predicted effective and safe concentration of 102 CFU/cm2 to 107 CFU/cm2.   
6.1.1.2.2. Preparation  
Boxed S. hominis A9 single-dose dispensers will be stored at -80°C until dispensation to the 
clinical staff.  S. hominis A9 will be thawed to 4°C prior to administration in clinic at the  
Treatment Visit (Day 0).  Further details regarding thawing and administration will be included in 
the Pharmacy Manual.  
6.1.1.2.3. Administration  
Participants will have a single dose of S. hominis A9 applied to one of their ventral arm (wrist to 
upper arm) by clinic staff.  The clinical staff member administering the product will record the 
associated dispenser ID and applicable arm.  
6.1.2. Placebo  
 
6.1.2.1.  
Formulation, Packaging, and Labeling   
Formulation of matching placebo will be identical to S. hominis A9, but without the bacteria and will be 
supplied in an identical sterile dispenser with spray head.  Each dispenser will contain 10 mL of placebo 
containing 85% USP grade PBS containing 15% USP grade glycerol.  Prior to shipping, each batch of 
placebo will undergo quality control testing, and additionally two aliquots from each lot will be 
randomly selected for release testing after packaging into sterile dispensers.  One aliquot will undergo 
Microbial Limits Testing in accordance with USP <61> and <62> testing, and the other will undergo visual 
testing.  If the product is not free from contaminants in accordance with USP <61> and <62> testing, the 
entire lot will be discarded appropriately, and will not be provided to the clinical site.  


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6.1.2.2.  
Dosage, Preparation, and Administration  
6.1.2.2.1. Dosage  
Each single-use dispenser will be used to dispense approximately 1 mL of placebo containing 85% 
PBS containing 15% glycerol.  To dispense approximately 1 mL, 6 pumps will be applied per dose 
(160 µL/pump).  
6.1.2.2.2. Preparation  
Boxed dispensers will be stored at -80°C until dispensation to the clinical staff.  Placebo will be 
thawed to 4 ̊C prior to administration in clinic at the Treatment Visit (Day 0).  Further details 
regarding thawing and administration will be included in the Pharmacy Manual.  
6.1.2.2.3. Administration  
Participants will have a single dose of placebo applied to their contralateral ventral arm (wrist to 
upper arm) by clinic staff.  The clinical staff member administering the product will record the 
associated dispenser ID and applicable arm.   
6.2. Investigational Product Accountability  
Under Title 21 of the Code of Federal Regulations (21CFR §312.62), the investigator will maintain adequate 
records of the disposition of the investigational product, including the date and quantity of the product 
received, to whom the product was dispensed (participant-by-participant accounting), and a detailed accounting 
of any product accidentally or deliberately destroyed.  Details of investigational product dispensation to the 
clinical site will be maintained by the manufacturing lab.   
Records for receipt, storage, use, and disposition will be maintained by the study site.  An investigational 
product-dispensing log will be kept current for each participant.  This log will contain the study identification 
number of each participant and the date and quantity of product dispensed.  
All records regarding the disposition of the investigational product will be available for inspection.  At the 
termination of the study, all dispensed, unused product will be destroyed.  For more information on handling of 
the investigational product, please refer to the Pharmacy Manual.  
6.3. Assessment of Participant Compliance with Investigational Agent  
Investigational product will be administered by a blinded study staff member and thus is an observed compliance.  
Participant compliance will be determined by the number and percentage of participants who receive a dose of ShA9 on 
their right or left arm and placebo on their contralateral arm at the Treatment Visit (Day 0).  Any deviations from the 
dosing schedule outside the defined visit windows will be recorded on the appropriate electronic case report form 
(eCRF).   
6.4. Toxicity Prevention and Management  
All steps will be taken to minimize potential risks of the study.  Participants with known sensitivities/allergies to any of 
the products used during this study will be excluded from study.  Participants with any cracks or breaks in their skin, 
including severely excoriated or bleeding skin suggesting that the patient may be susceptible to an infection, will be 
excluded.  Any AE, defined as any undesirable sign, symptom, or medical condition occurring after the participant’s 
written consent to participate is completed, will be recorded and reported to the Principal Investigator (PI), Division of 
Allergy, Immunology, and Transplantation (DAIT)/NIAID, FDA, and the IRB as required.  If the PI believes an AE is possibly 
related to the study product, the PI will determine whether or not it is in the best interest of the subject to continue in 


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the study.  If the participant is injured as a result of participation in this research, treatment will be available at the 
clinical center.  Further details of the services, including costs, and coverage will be explained in the informed consent, 
signed by the participant.  In the case of a skin infection, the participant would be treated according to standard 
treatment practices.  The treatment will include antibiotics (for infection) and Cetaphil® lotion (for irritation and 
dryness).  Refer to Section 7.4 for more information on rescue medications.  
6.5. Premature Discontinuation of Investigational Agent  
Study therapy may be prematurely discontinued, not applied fully to both the right and left ventral arms, for any 
participant who shows evidence of contact allergy to investigational product.  
Investigational product may also be prematurely discontinued for any participant if the investigator believes that 
continuing use of the investigational product is no longer in the best interest of the participant.  
If a subject is prematurely discontinued from study therapy, the subject will be asked to complete any remaining study 
visits, per protocol, through end of study (Day 31).  
7. Other Medications  
7.1. Concomitant Medications  
 
7.1.1  Protocol-mandated  
Participants will be required to apply Cetaphil® lotion to their ventral arms twice daily for approximately 
seven days (Pre-treatment Visit until Treatment Visit).  
 
7.1.2  Other permitted concomitant medications  
During study participation, participants are permitted to use topical steroids per discretion of the 
investigator on areas other than the ventral arms where treatment is applied and the designated 
swabbing area on the face.   
  
Use of antihistamines will not be exclusionary for protocol enrollment.  However, participants using 
antihistamines at the time of the Screening Visit will be asked to maintain a stable dose until completion 
of their last in-clinic visit.   
7.2. Prophylactic Medications  
Female participants of child-bearing potential must use an effective method of contraception (e.g. total abstinence, oral 
contraceptive, IUD, barrier method with spermicide, surgically sterilized partner, Depo-Provera, Norplant, NuvaRing, or 
hormonal implant) for the duration of study participation.   
Pregnancy tests will be performed on female participants of child-bearing potential who do not self-report as pregnant 
at the Screening and Treatment (Day 0) Visits.  At all other study visits, female participants of child-bearing potential will 
be asked if they are pregnant.   
All reported pregnancies in female participants and the partners of participants will be followed as described in Section  
12.6.  
7.3. Prohibited Medications/Therapies  
Participants are prohibited from using the following medications/therapies:  


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• 
Topical AD treatments (including steroids and topical calcineurin inhibitors) on the ventral arms where 
treatment is applied and the designated swabbing area on the face; Use of topical steroids on other areas is 
permitted per investigator discretion, as described in Section 7.1.2.  
• 
Any lotions, ointments or creams, including prescription moisturizers classified as a medical device (e.g., 
Atopiclair®, MimyX®, Epiceram®, etc.), other than Cetaphil® lotion (provided by the research clinic) on the 
ventral arms where treatment is applied and the designated swabbing area on the face; Use on other areas is 
permitted  
• 
Oral or topical antibiotics  
• 
Bleach baths  
• 
Oral or injectable AD therapies (steroids, immunosuppressive therapies, biologics)  
• 
Phototherapy (such as NBUVB, UVB, UVA1, PUVA) or tanning beds  
After the date of their CoNS falling below baseline density measured before application of ShA9 + 100 CFU/cm2, the 
participant will be allowed to use prohibited medications/therapies, as needed.  Use of medications/therapies will 
be recorded on the concomitant medications case report form (CRF).     
7.4. Rescue Medications   
If infection occurs, treatment will include topical and/or oral antibiotics depending on the severity of the infection 
clinically, and Cetaphil® lotion (for irritation and dryness).  The antibiotic sensitivity profile of Staphylococcus hominis A9, 
includes sensitivity to ampicillin-sulbactam, cefazolin, cefoxitin, clindamycin, daptomycin, levofloxacin, linezolid, 
minocycline, moxifloxacin, mupirocin, nitrofurantoin, oxacillin, rifampin, trimethoprim-sulfamethoxazole, and 
vancomycin; intermediate sensitivity to doxycycline; and resistance to erythromycin, penicillin G and tetracycline. The 
antibiotic of choice will depend on the individual participant’s medical history, including any possible contraindications 
to certain medications due to allergies or other concomitant medication use, but mupirocin will be the first line topical 
antibiotic of choice, and minocycline will be the first line oral antibiotic, followed by first generation cephalosporins, 
trimethoprim-sulfamethoxazole or rifampin.  Contact allergic reactions to Cetaphil lotion or glycerol will be treated with 
mid-potency topical steroids, such as fluocinonide ointment 0.05%.  
8. Study Procedures  
A summary of complete study procedures is included in Appendix B Schedule of Events.  To ensure participant safety, 
participants will be asked to comply with UCSD’s current institutional policies and procedures related to COVID-19, 
which are based on recommendations from the Centers for Disease Control and Prevention when scheduling and 
attending their appointments.  
8.1. Recruitment Visit  
Potential participants will be recruited using standardized questionnaires that collect contact information and disease 
status related to inclusion and exclusion criteria.  Participants may be recruited by phone or in person. Potential 
participants from UCSD departments outside of Dermatology will be identified using EPIC slicer dicer tool, then 
confirmed with their primary care provider to see if their patients can be approached for recruitment. Once confirmed 
by their primary care provider, patients will be contacted through a message on MyChart to assess their interest in 
participating. Those interested will then be followed up by a phone call to assess their eligibility. Those who have no 
obvious characteristics making them ineligible for the study and who are interested in participating will be invited to 
clinic to complete the Screening Visit.   


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8.2. Screening Visit (Day -14 to -7)  
The purpose of the Screening Visit is to confirm eligibility to continue in the study.  During the visit, written informed 
consent will be obtained from the participant prior to performing any other study procedures.   
The following procedures and assessments will be conducted to determine participant eligibility:   
• 
Collection of demographics  
• 
Medical History and physical examination, including assessment of AD severity (IGA, L-EASI on the 
ventral arms, SCORAD, and local pruritus NRS) by a study physician or other qualified medical 
professional  
• 
Pregnancy status and urine pregnancy test: Female participants will be asked if they are pregnant.  A 
urine pregnancy test will be completed for all female participants of child bearing potential who do not 
self-report as pregnant.  
• 
Assessment of concomitant medications  
• 
Vital signs and growth parameters (height and weight)  
• 
Assessment of Adverse Events (AEs)  
Participants who meet inclusion and exclusion criteria including the required washout periods for medications/therapies 
will be considered screened and eligible.  Participants who do not meet inclusion and exclusion criteria due to 
assessment of their concomitant medications and require a washout of 14 days or more will be required to rescreen, 
prior to enrolling in the study.  Participants who do not wish to washout of the prohibited medications or do not meet 
full inclusion and exclusion criteria will be identified as screen failures and will not continue in the study.   
8.3. Pre-treatment Visit (Day -7 ± 1 Day)  
Screened and eligible participants will be invited to complete the Pre-treatment Visit.  If a participant meets all required 
washout periods for medications/therapies during their Screening Visit, they may complete the Pre-treatment Visit on 
the same day of the Screening Visit.  The Pre-treatment Visit must be completed 7 days (± 1 Day) prior to the Day 0 Visit.  
During this visit, participants will have their initial skin swabs collected for the assessment of live bacteria by culture (S. 
aureus and CoNS) and bacterial DNA (S. hominis A9) by qPCR.   
The following procedures, assessments, and laboratory measures will be conducted at this Pre-treatment Visit:   
• 
Interim medical history and physical exam, including assessment of AD severity (L-EASI, SCORAD, and 
local Pruritus NRS) by study physician or other qualified medical professional (if conducted on a different 
day than the Screening Visit)  
• 
Pregnancy status of participant and partner of participant, as applicable (if conducted on a different day 
than the Screening Visit)   
• 
Assessment of concomitant medications (if conducted on a different day than the Screening Visit)  
• 
Vital signs (if conducted on a different day than the Screening Visit)  
• 
Identification of measured lesional and non-lesional sites on the participant’s ventral arms, including 
photographs, and skin swab collection, to assess live bacteria by CFU and bacterial DNA by qPCR.  Two 
lesional and two non-lesional swabs will be collected from each arm (8 swabs total).    
• 
Assessment of AEs  
During the Pre-treatment Visit, participants will be given Cetaphil® lotion and Dove moisturizing soap.  Participants will 
be instructed to apply a thin layer of Cetaphil® lotion to each arm, twice a day for approximately 1 week (Pre-treatment 


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Visit until Treatment Visit) and to use the soap when they shower, avoiding the arms.  At the conclusion of the visit, 
participants will be given instructions regarding restrictions on bathing/showering, exercise, and use of emollient.  
Participants will be instructed to refrain from swimming in chlorinated pools and hot tubs and to refrain from the use of 
prohibited medications, for the duration of their study participation.  Female subjects of child-bearing potential will be 
instructed to use an effective method of birth control or abstinence.   
8.4. Treatment Visit (Day 0)  
Prior to the Treatment Visit, the clinical team will contact study participants to confirm if their pre-treatment swab 
results meet criteria for enrolling in the AD SA+ or AD SA- group.  If the enrollment goal(s) for AD SA+ or AD SA- have 
been met, the participant will be identified as a screen failure.   
The purpose of the Day 0 Treatment Visit is to complete study treatment, application of ShA9 and placebo.  The 
following procedures, assessments, and laboratory measures will be conducted at the Treatment Visit:  
• 
Interim medical history and physical exam, including assessment of AD severity (L-EASI, SCORAD, and 
local Pruritus NRS) by study physician or other qualified medical professional  
• 
Pregnancy status of participant and partner of participant, as applicable, and urine pregnancy test 
(female participants only), per Section 8.2  
• 
Assessment of concomitant medications  
• 
Vital Signs  
• 
Randomization   
• 
Identification of measured non-lesional site on the participant’s face, including photograph, and skin 
swab collection for the assessment of ShA9 spread.  Two non-lesional swabs will be collected from the 
participant’s face.  
• 
Skin Swab Collections from the ventral arms, to assess live bacteria by CFU, bacterial DNA by qPCR, and 
antibiotic sensitivity to penicillin G, tetracycline, and erythromycin.  Two lesional and two non-lesional 
swabs will be collected from each ventral arm (8 swabs total).  
• 
S. hominis A9 and placebo application  
• 
Participants will remain in clinic for up to 6 hours following the application of investigational product.  
Additional skin swabs will be collected from the participant’s arms at 15 minutes, 1, 2, 4, and 6 hours 
post-application.  Two lesional and two non-lesional swabs will be collected from each ventral arm for a 
total of eight swabs at each time point.    
• 
Assessment of AEs  
At the conclusion of this visit, participants will be reminded to use Dove moisturizing soap, whenever they shower, but 
to avoid the arms, until after their last in-clinic visit is complete.  Participants will be allowed to use Cetaphil® lotion to 
moisturize if necessary.  Participants will be given instructions regarding restrictions on bathing/showering, exercise, and 
use of emollient.   Participants will be reminded to refrain from swimming in chlorinated pools and hot tubs and to 
refrain from the use of prohibited medications, for the duration of their study participation.  Participants will be asked to 
try to restrict pets from licking the treated areas.  Female subjects of child-bearing potential will be instructed to use an 
effective method of birth control or abstinence.  
Participants will be given a handcard describing the symptoms of skin and systemic infections and will be told to contact 
the research clinic if they experience any of the listed symptoms.  The participants will be instructed to take the 
handcard with them should they seek medical care at another facility.  The handcard will state that the patient is 


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currently in a study involving the application of non-pathogenic bacteria to the skin and will instruct them to call the 
research site if an infection of the skin or blood is suspected.  
8.5. Follow-Up Visits (24 Hour and Days 3, 10, 17 and 24)   
After the Treatment Visit, participants will be asked to return to the clinic for a follow up 24 Hours after the Day 0 Visit 
and on Days 3 (± 1 day), 10 (± 1 day), 17 (± 2 days), and 24 (± 2 days).  The purpose of the follow-up visits is to assess S. 
aureus and CoNS bacteria abundance and AEs.  
The following procedures, assessments, and laboratory measures will be conducted at each visit:  
• 
Assessment of AEs  
• 
Assessment of AD severity (L-EASI, SCORAD, and local Pruritus NRS) by study physician or other qualified 
medical professional   
• 
Pregnancy status of participant and partner of participant, as applicable  
• 
Assessment of concomitant medications  
• 
Vital Signs  
• 
Skin Swab Collections, (lesional and non-lesional) from the ventral arms, to assess live bacteria by CFU 
and bacterial DNA by qPCR and non-lesional skin swab collection from the participant’s face for the 
assessment of ShA9 spread.  Two lesional and two non-lesional swabs will be collected from each 
ventral arm, and two additional non-lesional swabs will be collected from the face (10 swabs total).  
• 
Lesional skin swabs taken at the last Follow-Up Visit will also be analyzed for antibiotic sensitivity to 
penicillin G, tetracycline, and erythromycin.   
At the conclusion of each visit, participants will be reminded to use Dove moisturizing soap, whenever they shower, but 
to avoid the arms, until after their last in-clinic visit is complete.  Participants will be given instructions regarding 
restrictions on bathing/showering, exercise, and use of emollient.  Participants will be reminded to refrain from 
swimming in chlorinated pools and to refrain from the use of prohibited medications.  Female participants of 
childbearing potential will be instructed to use an effective method of birth control or abstinence.   
Upon completion of the Day 3 Visit, a participant will be notified prior to their next scheduled follow-up visit if they need 
to return to the clinic.  A participant will not be required to complete the remaining follow up clinic visits (Day 10 
through Day 24) if the lesional swabs from both arms collected at their most recent prior visit are CoNS negative, 
defined as below baseline density measured before application of ShA9 + 100 CFU/cm2. The lesional skin swabs taken at 
the last completed Follow-Up Visit will be used in analysis for antibiotic sensitivity to penicillin G, tetracycline, and 
erythromycin. Once a participant completes his/her required follow up clinic visits, he/she may resume prohibited 
medications as described in Section 7.3, as needed, and may use his/her choice of soap and emollient.  
8.6. End of Study Phone Visit (Day 31 ± 7 Days)  
The purpose of the Day 31 End of Study Phone Visit is to complete participant follow-up.  This visit will be brief, and 
participants will be asked to report any new AEs and to answer questions regarding the status of their AD and 
concomitant medication use.  Female participants of child-bearing potential will be asked whether they have tested 
positive to a pregnancy test since their last visit. Participants will be asked whether their partner has tested positive to a 
pregnancy test since their last visit, as applicable.   
Any participant with an ongoing AE/Serious Adverse Event (SAE) at the time of this visit will continue to be followed until 
the event is resolved with or without sequelae or the AE/SAE stabilizes.   


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8.7. Unscheduled Visits   
If disease activity increases, participants experience signs and symptoms as described on the study handcard, or other 
concerns arise between regularly scheduled visits, participants will be instructed to contact study personnel and may be 
asked to return to the study site for an “unscheduled” visit.   
The following procedures, assessments, and laboratory measures will be conducted at the Unscheduled Visit:  
• 
Assessment of AEs  
• 
Physical examination, including assessment of AD severity (L-EASI, SCORAD, and local Pruritus NRS) by 
study physician or other qualified medical professional   
• 
Pregnancy status of participant and partner of participant, as applicable  
• 
Vital Signs  
• 
Skin Swab collection, per investigator discretion  
• 
Blood collection, per investigator discretion  
If a participant presents to the research clinic with a suspected skin or systemic infection, skin swabs and/or blood 
samples may be collected.  If collected, skin swab samples will be cultured and speciated to determine whether the 
transplant Staph strain caused the infection.  A complete blood count with differential and blood culture will be 
performed on the blood sample, if collected, to assess for infection.   
Participants will be given a handcard to present to clinical staff should they seek care for an infection at another facility.  
The handcard, as described in Section 8.4, will state that the participant is currently in a clinical trial involving the 
application of non-pathogenic bacteria to the skin, list the sensitivities of S. hominis, and include instructions to call the 
research site if an infection of the skin or blood is suspected.   
8.8. Visit Windows  
Study visits should take place within the time limits specified above: the designated visit windows (i.e. +/- n days) for 
each scheduled visit are also indicated in Appendix B Schedule of Events.   
9. Mechanistic Assays  
This section describes the proposed methodologies for this study.  The techniques are state-of-the-art at the time of the 
writing of this protocol.  Even so, the techniques will be updated and/or changed should there be additional technical 
breakthroughs in this area of research.  Details of the laboratory processes are described in Standard Operating 
Procedures (SOPs) maintained by each laboratory.   
9.1. Skin Swab Collection Procedure  
Lesional and non-lesional swabbing areas (21 cm2 total for each; 7 areas of 3 cm x 1 cm of lesional and non-lesional skin 
each) will be defined on each ventral arm at the Pre-treatment Visit.  The lesional swab locations will be selected from 
the most severe lesional areas and may not be contiguous.  Lesions closer to the antecubital region will take precedence 
in defining these areas.  The non-lesional swab locations will be defined as areas of normal skin on the ventral arms and 
also may not be contiguous.  In addition, a non-lesional swabbing area of 3 cm x 1 cm will be defined on the face at the 
Treatment Visit.  The designated lesional and non-lesional sites for swabbing will be marked with a pen and 
nonidentifying digital photographs will be taken for reference during future visits.  The number and location of swabs to 
be collected at each visit and time point are delineated in Section 8 and the Schedule of Events.  Each swab will cover a 3 
cm2 area.  The lateral edge of the swab will be rubbed across the measured area in a cross-wise manner while rotating 


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the swab handle between the thumb and forefinger.  Swabbing will be performed for at least 30 seconds.  Refer to the 
MOP for additional details regarding the collection of skin swabs.   
9.2. CFU Quantification of Live Staphylococcus aureus and CoNS  
Skin swabs will be collected from pre-measured areas of lesional and non-lesional skin per Section 9.1 using swabs 
premoistened with TSB.  Collected swab tips for CFU quantification will be stored in 1 mL of 85% TSB/15% Glycerol and 
stored at -80oC until analysis.  Live CoNS will be measured by counting CFU on mannitol salt agar with egg yolk, which is 
a selective media for staphylococcus species.  CoNS and S. aureus can be distinguished by the colony morphology (CoNS: 
pink color without egg yolk reaction; S. aureus: yellow color with egg yolk reaction).  Live S. aureus will be measured on 
Bear-Parker agar, which is a selective media for S. aureus.  CFU quantification will give us quantification of live S. aureus 
and CoNS, as each will be grown in selective media and are important for our Primary Endpoint, Secondary Endpoint 1, 
and Exploratory Endpoints 2-5.   
9.3. Quantification of S. hominis A9 CFU   
To specifically identify colonies of S. hominis A9, 24-48 CoNS colonies will be picked from each culture plate and 
subjected to colony PCR using strain specific primers for the lantibiotic gene.  CFUs of S. hominis A9 will be estimated 
from total CoNS CFU × % of S. hominis A9 positive colonies.  Quantification of S. hominis A9 estimated CFU will allow us 
to quantify S. hominis A9 specifically as outlined in Exploratory Endpoint 1.    
9.4. Quantification of Bacterial DNA  
Skin swabs will be collected from pre-measured areas of lesional and non-lesional skin per Section 9.1, using swabs 
premoistened with Tris-EDTA buffer containing 0.1% TritonX-100 and 0.05% Tween-20 (w/v).  Microbial DNA will be 
extracted with the PureLink Microbiome DNA Purification kit (Invitrogen).  Abundance of DNA from total staphylococcus 
(G-staph), S. aureus, S. hominis and Sh-lantibiotic will be quantified by qPCR using genus-, species- and gene- specific 
primer sets, respectively.  Quantification of bacterial DNA will allow for the assessment of specific shifts in the 
microbiome pre and post application of S. hominis A9, correlation between S. hominis A9 and Sh-lantibiotic, and 
correlation between S. aureus and Sh-lantibiotic, supporting the primary objective to assess the factors that overall 
influence survival.    
9.5 Analysis of Antibiotic Sensitivity  
Lesional skin swabs from the Baseline Visit and last Follow-Up Visit will be tested for antimicrobial sensitivity to 
erythromycin, tetracycline, and penicillin G. The same volume of bacterial swab samples from lesional skin at Baseline 
and last Follow-Up Visit time points will be spread on culture plates containing Mannitol-salt agar, a staphylococcus 
selective agar, without or with erythromycin (4 µg/mL), or tetracycline (8 µg/mL), or penicillin G (1 µg/mL). The 
concentration of each antibiotic was determined based on the minimal inhibitory concentration used to define ShA9 as 
resistant to these antibiotics. The proportion of CoNS with antibiotic sensitivity will be determined by comparing the 
CFU measured on duplicate agar plates without antibiotic to the CFU measured on agar plates with each antibiotic by 
the equation [(CFU without antibiotic containing plate - CFU with antibiotic containing plate) / CFU without antibiotic 
containing plate]. The proportion of antibiotic-sensitive CoNS detected at baseline will be compared to the proportion 
from swabs collected at the last Follow-Up Visit. If an increase in the proportion of antibiotic resistant CoNS is measured 
to be greater than 10%, resistant colonies will be evaluated to determine if they are ShA9. If these colonies are not ShA9, 
DNA sequencing will be performed to determine if they contain antibiotic resistance genes that were originally present 
in ShA9, thus indicating the likelihood for horizontal gene transfer.  


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10. Biospecimen Storage  
Lesional and non-lesional skin swabs will be obtained from all study participants.  These skin swabs and any derivative 
samples will be obtained for potential future analysis of additional parameters that describe the composition and 
function of the skin microbiome.  Participants will be asked to give permission for long-term storage and future use 
during the consent process.  All samples stored for future use will be kept in a central repository at UCSD.   
Instructions for sample preparation, handling, storage, and shipping are included in the MOP.  The PIs will be responsible 
for acknowledging and implementing all the regulations for classification, sample handling, packaging and labeling, 
permits or authorizations, and personnel training for shipment of biological and hazardous materials required for the 
conduct of this study.  
11. Criteria for Participant and Study Completion and Premature Study Termination  
11.1.  
Participant Completion  
Participant participation will be defined as complete at the conclusion of the Day 31 End of Study Phone Visit.   
11.2.  
Participant Stopping Rules and Withdrawal Criteria  
Participants may be prematurely terminated from the study for the following reasons:  
1. The participant elects to withdraw consent from all future study activities, including follow-up.  
2. The participant is “lost to follow-up” (i.e., no further follow-up is possible because attempts to reestablish 
contact with the participant have failed).   
3. The participant dies.   
4. The Investigator no longer believes participation is in the best interest of the participant.  
5. Individual safety stopping rules   
a. Evidence of infection either cutaneously or systemically which in the opinion of the investigator makes 
the participant a poor candidate for receipt of the investigational product or requires treatment with a 
prohibited medication/therapy that would interfere with assessment of the study endpoints.  
b. Severe worsening of AD which in the opinion of the investigator makes the participant a poor candidate 
for receipt of the investigational product or requires treatment with a prohibited medication/therapy 
that would interfere with assessment of the study endpoints.  
c. Pregnancy in a female participant prior to the Treatment Visit (Refer to Section 12.6).   
11.3.  
Participant Replacement  
If any participants withdraw or are withdrawn prior to achieving CoNS elimination (CoNS CFU below baseline density 
measured before application of ShA9 + 100 CFU/cm2), additional participants may be enrolled as necessary to obtain a 
modified ITT sample as defined in Section 13.4.1.   
11.4.  
Follow-up after Early Study Withdrawal  
If a participant is withdrawn from the study for any reason, the participant will be asked to complete a final visit by 
phone to assess any AEs since his/her last visit and to answer questions about the status of his/her AD.  Any participant 
with an ongoing AE/SAE at the time of this phone contact will continue to be followed until the event is resolved with or 
without sequelae or until the AE stabilizes or until 30 days after the participant prematurely withdraws (without 
withdrawing consent)/or is withdrawn from the study, whichever occurs first.  
  


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11.5.  
Study Stopping Rules  
Screening, enrollment, and application of treatment for this trial will be suspended pending immediate DSMB review for 
the following reasons:  
• 
A single participant experiences any SAE for which there is a reasonable possibility that the investigational 
product caused the SAE  
• 
The development of any severe (Grade 3) AE for which attribution is defined as related or possibly related   
Follow up visits will continue to be conducted per protocol while under DSMB review.  After review of the data, the 
DSMB will make recommendations regarding study conduct and/or continuation.  
12. Safety Monitoring and Reporting  
12.1 Overview  
This section defines the types of safety data that will be collected under this protocol and outlines the procedures for 
appropriately collecting, grading, recording, and reporting those data.  AEs that are classified as serious according to the 
definition of health authorities must be reported promptly (per Section 12.5) to the sponsor DAIT/NIAID.  Appropriate 
notifications will also be made to site PIs, IRB, and health authorities.   
Information in this section complies with International Conference on Harmonisation (ICH) Guideline E2A: Clinical Safety  
Data Management: Definitions and Standards for Expedited Reporting, ICH Guideline E-6: Guideline for Good Clinical  
Practice (GCP), 21CFR Parts 312 and 320, and applies the standards set forth in the National Cancer Institute (NCI), 
Common Terminology Criteria for Adverse Events (CTCAE), Version 5.0: http://ctep.cancer.gov/reporting/ctc.html.  
12.2 Definitions  
12.2.1 Adverse Event (AE)  
Any untoward or unfavorable medical occurrence associated with the subject’s participation in the research, 
whether or not considered related to the subject’s participation in the research (modified from the definition of 
adverse events in 21 CFR 312.32(a), the International Council for Harmonisation of Technical Requirements for 
Pharmaceuticals for Human Use (ICH) E-6(R2) Guidelines for Good Clinical Practice and OHRP "Guidance on 
Reviewing and Reporting Unanticipated Problems Involving Risks to Subjects or Others and Adverse Events 
(1/15/07)" http://www.hhs.gov/ohrp/policy/advevntguid.html#Q2 ).   
.  
For this study, an AE will include any untoward or unfavorable medical occurrence associated with:   
• 
Study Treatment regimen: An AE occurring from the time of consent and within one month of  
S. hominis A9 product administration   
• 
Study mandated procedures:   
o Blood Draw (to only occur at Unscheduled Visit at the discretion of the study physician 
or other qualified medical professional)  
The following events related to the blood draw procedure will be considered an AE if they 
occur within 48 hours of the blood draw:  
o Fainting / Vasovagal Events  


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o Bruising at the puncture site larger than 2 cm in diameter o Bleeding from the puncture 
site lasting more than 30 minutes o Swelling at puncture site larger than 2 cm  
12.2.1.1 Suspected Adverse Reaction (SAR)   
Any adverse event for which there is a reasonable possibility that the drug caused the adverse event.  
For the purposes of IND safety reporting, ‘reasonable possibility’ means there is evidence to suggest a 
causal relationship between the drug and the adverse event.  Suspected adverse reaction implies a 
lesser degree of certainty about causality than adverse reaction, which means any adverse event caused 
by a drug (21 CFR 312.32(a)).  
12.2.2 Unexpected Adverse Event or Unexpected Suspected Adverse Reaction  
An adverse event or suspected adverse reaction is considered “unexpected” if it is not listed in the Investigator 
Brochure or is not listed at the specificity or severity that has been observed; or, if an investigator brochure is 
not required or available, is not consistent with the risk information described in the general investigational plan 
or elsewhere in the current application, as amended.  For example, under this definition, hepatic necrosis would 
be unexpected (by virtue of greater severity) if the investigator brochure referred only to elevated hepatic 
enzymes or hepatitis.  Similarly, cerebral thromboembolism and cerebral vasculitis would be unexpected (by 
virtue of greater specificity) if the investigator brochure listed only cerebral vascular accidents.  "Unexpected," 
as used in this definition, also refers to adverse events or suspected adverse reactions that are mentioned in the 
investigator brochure as occurring with a class of drugs or as anticipated from the pharmacological properties of 
the drug, but are not specifically mentioned as occurring with the particular drug under investigation 21 CFR 
312.32(a)).  
12.2.3 Serious Adverse Event (SAE) or Serious Suspected Adverse Reaction  
An adverse event or suspected adverse reaction is considered “serious” if, in the view of either the investigator 
or the Sponsor, it results in any of the following outcomes: Death, a life-threatening adverse event, inpatient 
hospitalization or prolongation of existing hospitalization, a persistent or significant incapacity or substantial 
disruption of the ability to conduct normal life functions, or a congenital anomaly/birth defect.  Important 
medical events that may not result in death, be life-threatening, or require hospitalization may be considered 
serious when, based upon appropriate medical judgment, they may jeopardize the patient or subject and may 
require medical or surgical intervention to prevent one of the outcomes listed in this definition.  Examples of 
such medical events include allergic bronchospasm requiring intensive treatment in an emergency room or at 
home, blood dyscrasias or convulsions that do not result in inpatient hospitalization, or the development of drug 
dependency or drug abuse  (21 CFR 312.32(a)).  
Elective hospitalizations are not to be reported as an SAE unless hospitalization is prolonged due to 
complications.  
12.3 Grading and Attribution of Adverse Events  
12.3.1 Grading Criteria  
The study site will grade the severity of AEs experienced by the study participants according to the criteria set 
forth in the NCI-CTCAE Version 5.0.  This document (referred to herein as the NCI-CTCAE manual) provides a 
common language to describe levels of severity, to analyze and interpret data, and to articulate the clinical 
significance of all AEs.  The NCI-CTCAE has been reviewed by the study investigators and sponsor and has been 
deemed appropriate for the participant population to be studied in this protocol.   


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AEs will be graded on a scale from 1 to 5 according to the following standards in the NCI-CTCAE manual:  
Grade 1 = mild AE  
Grade 2 = moderate AE  
Grade 3 = severe AE  
Grade 4 = life-threatening AE or urgent intervention indicated  
Grade 5 = death  
Events grade 1 or higher will be recorded on the appropriate AE eCRF for this study.  
For grading an abnormal value or result of a clinical or laboratory evaluation (including, but not limited to, a 
radiograph, an ultrasound, an electrocardiogram etc.), a treatment-emergent AE is defined as an increase in 
grade from baseline or from the last post-baseline value that doesn’t meet grading criteria.  Changes in grade 
from screening to pre-treatment and enrollment at Day 0 will also be recorded as AEs, but are not 
treatmentemergent.  If a specific event or result from a given clinical or laboratory evaluation is not included in 
the NCICTCAE manual, then an abnormal result would be considered an AE if changes in therapy or monitoring 
are implemented as a result of the event/result.  
For additional information and a printable version of the NCI-CTCAE manual, consult the NCI-CTCAE web site:  
http://ctep.cancer.gov/reporting/ctc.html.  
12.3.2 Attribution Definitions  
The relationship, or attribution, of an AE to the investigational product regimen or study procedure(s) will 
initially be determined by the site investigator and recorded on the appropriate AE/SAE eCRF.  Final 
determination of attribution for safety reporting will be determined by the DAIT/NIAID Medical Monitor.  The 
relationship of an AE to study therapy regimen or procedures will be determined using the descriptors and 
definitions provided in Table 12.3.2.   
        
Table 12.3.2 Attribution of Adverse Events  
Code  
Descriptor  
Relationship (to investigational product regimen or 
study procedure: blood draw)  
UNRELATED CATEGORY  
 
1  
Not  
Related  
The adverse event is clearly not related:  there is 
insufficient evidence to suggest a causal relationship.  
 RELATED CATEGORIES  
 
2  
Possible  
The adverse event has a reasonable possibility to be 
related; there is evidence to suggest a causal 
relationship.  
3  
Related  
The adverse event is clearly related.  


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12.4 Collection and Recording of Adverse Events  
12.4.1 Collection Period  
AEs (including SAEs) will be collected from the time of consent until a participant completes study participation 
or until 30 days after he/she prematurely withdraws (without withdrawing consent) or is withdrawn from the 
study.  
12.4.2 Collecting Adverse Events  
AEs (including SAEs) may be discovered through any of these methods:  
• 
Observing the participant  
• 
Interviewing the participant in an objective manner [e.g., using structured questioning]  
• 
Receiving an unsolicited complaint from the participant  
• 
Receiving a call from the participant outside of their regular study visits; Instructions for contacting the clinic 
will be provided on an instructional handcard at the Treatment Visit.  
• 
In addition, an abnormal value or result from a clinical or laboratory evaluation can also indicate an AE, as 
defined in Section 12.3.  
12.4.3 Recording Adverse Events  
Throughout the study, the investigator will record AEs and SAEs as described previously (Section 12.2) on the 
appropriate AE/SAE eCRF regardless of the relationship to study therapy regimen or study procedure.  
Once recorded, an AE/SAE will be followed until it resolves with or without sequelae or until the end of study 
participation, or until 30 days after the participant prematurely withdraws (without withdrawing consent)/or is 
withdrawn from the study, whichever occurs first.  
12.5 Reporting of Serious Adverse Events and Adverse Events  
12.5.1 Reporting of Serious Adverse Events to DAIT/NIAID   
This section describes the responsibilities of the site investigator to report SAEs to the sponsor and the SACCC 
via the SAE eCRF.  Timely reporting of AEs is required by 21 CFR 312.32 and ICH E6 guidelines.   
Site investigators will report all SAEs (see Section 12.2.3), regardless of relationship or expectedness within 24 
hours of discovering the event.  
For SAEs, all requested information on the SAE eCRF will be provided.  However, unavailable details of the event 
will not delay submission of the known information.  An Initial SAE eCRF shall include as much information as 
possible, but at a minimum will include the following:   
• 
AE term   
• 
Relationship to investigational product  
• 
Relationship to stopping protocol prohibited medications/therapy  
• 
Relationship to study procedure   
• 
Reason why the event is serious  


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• 
Supplementary CRF pages that are current at the time of SAE reporting: medical history, 
concomitant medications, demographics, and investigational product administration  
As additional details become available, the SAE eCRF should be updated and submitted.  Every time the SAE 
eCRF is submitted, it should be electronically signed by the investigator or sub-investigator.   
  
For additional information regarding SAE reporting, contact Rho Product Safety:   
Rho Product Safety   
2635 E. NC Hwy 54  
Durham, NC 27713  
Toll-free: 1-888-746-7231   
SAE Fax Line: 1-888-746-3293   
Email: rho_productsafety@rhoworld.com  
12.5.2 Reporting to Health Authority  
After an AE, requiring 24-hour reporting (per Section 12.5.1), is submitted by the site investigator and assessed 
by DAIT/NIAID, there are two options for DAIT/NIAID to report the AE to the appropriate health authorities:  
12.5.2.1 Annual Reporting  
DAIT/NIAID will include in the IND Annual Report to the FDA all reported AEs classified as:   
o Serious, expected, suspected adverse reactions (see Section 12.2.1.1 and Section 12.2.2) 
o Serious and not a suspected adverse reaction (see Section 12.2.2) o Pregnancies  
Note that all AEs (not just those requiring 24-hour reporting) will be reported in the IND Annual Report.  
12.5.2.2 Expedited Safety Reporting   
This option, with 2 possible categories, applies if the adverse event is classified as one of the following:  
Category 1:  Serious and unexpected suspected adverse reaction [SUSAR] (see Section 12.2.1.1, Section  
12.2, and 21 CFR 312.32(c) (1) i).  
The sponsor shall report any suspected adverse reaction that is both serious and unexpected.   
The sponsor shall report an adverse event as a suspected adverse reaction only if there is  
evidence to suggest a causal relationship between the investigational product and the adverse 
event, such as:  
1. A single occurrence of an event that is uncommon and known to be strongly associated 
with investigational product exposure (e.g., sepsis, cellulitis, or deep tissue infections);  
2. One or more occurrences of an event that is not commonly associated with 
investigational product exposure, but is otherwise uncommon in the population 
exposed to the investigational product (e.g., tendon rupture);  
3. An aggregate analysis of specific events observed in a clinical trial (such as known 
consequences of the underlying disease or condition under investigation or other events 
that commonly occur in the study population independent of investigational product 
regimen) that indicates those events occur more frequently in participants receiving the 
investigational product than in a concurrent or historical control group.  


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Category 2: Any findings from studies that suggests a significant human risk   
The sponsor shall report any findings from other epidemiological studies, analyses of adverse 
events within the current study or pooled analysis across clinical studies, or animal or in vitro 
testing (e.g. mutagenicity, teratogenicity, carcinogenicity) that suggest a significant risk in 
humans exposed to the investigational product that would result in a safety-related change in 
the protocol, informed consent, investigator brochure or other aspects of the overall conduct of 
the study.    
DAIT/NIAID shall notify the FDA and all participating investigators of expedited Safety Reports within 15 
calendar days; unexpected fatal or immediately life-threatening suspected adverse reaction(s) shall be 
reported as soon as possible or within 7 calendar days.  
12.5.3 Reporting of Adverse Events to IRB/IEC  
All investigators shall report AEs, including expedited reports, in a timely fashion to their local IRB in 
accordance with applicable regulations and guidelines.  All Safety Reports to the FDA shall be distributed by 
DAIT/NIAID or designee to UCSD for site IRB submission.  
12.6 Pregnancy Reporting  
The investigator shall be informed of any pregnancy in a female study participant immediately upon becoming 
aware of the event.  If a female participant becomes pregnant prior to the Treatment Visit, the participant will 
no longer be eligible to participate in the study and will be withdrawn as described in Section 11.2.   If a 
pregnancy occurs following the Treatment Visit, the pregnant participant will continue to be followed per the 
Schedule of Events.  The investigator shall counsel the participant and discuss the risks of continuing with the 
pregnancy and the possible effects on the fetus.  Monitoring of the pregnant participant shall continue until the 
conclusion of the pregnancy.    
The investigator shall report to the SACCC all pregnancies within 1 business day of becoming aware of the event 
using the Pregnancy eCRF.  The SACCC will report all pregnancies to DAIT/NIAID using the procedures as outlined 
in the Safety Management Plan.   All pregnancies identified during the study shall be followed to conclusion, and 
the outcome of each must be reported.  The Pregnancy eCRF shall be updated and submitted to the SACCC  
when details about the outcome are available.  When possible, similar information shall be obtained for a 
pregnancy occurring in a partner of a study participant.  Information on this pregnancy will be collected on the 
Pregnancy eCRF.  In order for Sponsor or designee to collect any pregnancy surveillance information from the 
female partner, the female partner must sign an informed consent form for disclosure of this information.  
Information requested about the delivery shall include:  
o 
Gestational age at delivery o  
Birth weight, length, and head 
circumference o  
Gender  
o 
Appearance, pulse, grimace, activity, and respiration (APGAR) score at 1 
minute, 5 minutes, and 24 hours after birth, if available o  
Any 
abnormalities.  


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Should pregnancy complications result in a congenital abnormality, birth defect, miscarriage, or medically 
indicated abortion - an SAE shall be submitted to the SACCC using the SAE reporting procedures described 
above.  
12.7 Reporting of Other Safety Information  
An investigator shall promptly notify their local IRB, in accordance with applicable regulations and guidelines, as 
well as the SACCC and DAIT/NIAID via email when an “unanticipated problem involving risks to participants or 
others” is identified, which is not otherwise reportable as an adverse event.  
12.8 Review of Safety Information  
12.8.1 Medical Monitor Review  
The DAIT/NIAID Medical Monitor shall receive monthly reports from the SACCC compiling new and accumulating 
information on AEs, SAEs, and pregnancies recorded by the study site on appropriate eCRFs.  
In addition, the DAIT/NIAID Medical Monitor shall review and make decisions on the disposition of the SAE and 
pregnancy reports received by the SACCC (See Sections 12.5.1 and 12.6).   
12.8.2 DSMB Review   
The SACCC will provide the NIAID Allergy-Asthma [Alpha] DSMB with a listing of all AEs and SAEs at least 
annually.  Furthermore, the DSMB will be informed of expedited reports of SAEs.  
12.8.2.1 Planned DSMB Reviews   
The DSMB shall review safety data at least yearly during planned DSMB Data Review Meetings.  Data for 
the planned safety reviews will include, at a minimum, a listing of all reported AEs and SAEs.    
12.8.2.2 Ad hoc DSMB Reviews   
In addition to the pre-scheduled data reviews and planned safety monitoring, the DSMB may be called 
upon for ad hoc reviews when an event occurs that is of sufficient concern to the DAIT/NIAID Medical 
Monitor and/or the protocol chair(s) to warrant DSMB review, this includes all Expedited Safety Reports.  
The DSMB will be notified within 24 to 48 hours by the NIAID Medical Monitor and will promptly review 
any event that potentially impacts safety at the request of the protocol chair or DAIT/NIAID or any 
occurrence that meets the definition of the Study Stopping Rules defined in Section 11.5.   
After review of the data, the DSMB will make recommendations regarding study conduct and/or 
continuation.  
12.8.2.2.1 Temporary Suspension of Enrollment and Study Treatment for ad hoc DSMB Safety 
Review   
A temporary halt in screening, enrollment, and investigational product application will be 
implemented if an ad hoc DSMB safety review is required.  New participants will not be 
consented for study participation during the enrollment and treatment halt.  Participants who 
were screened or who completed the Pre-treatment Visit but who are not yet randomized will 
not be allowed to continue with the Day 0 Treatment Visit.  All participants not randomized 
within 14 days of the Screening Visit must rescreen.  Follow up visits will continue per the 
Schedule of Events.   


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13. Statistical Considerations and Analytical Plan  
13.1.  
Overview   
This is a pilot, open-label, single-center trial designed to assess the survival of transplanted S. hominis A9 on the skin of 
adults with moderate-to-severe atopic dermatitis on the ventral arms who are culture positive or negative for S. aureus 
colonization.  
13.2.  
Endpoints  
13.2.1 Primary Endpoint   
The primary endpoint for this pilot study is the duration of survival, measured as the time needed for the CoNS 
CFU to drop below baseline density measured before application of ShA9 + 100 CFU/cm2 on the lesional ventral 
arm skin of AD participants positive for S. aureus (AD SA+)  
13.2.2 Secondary Endpoints  
1. The duration of survival, measured as the time needed for the CoNS CFU to drop below baseline density 
measured before application of ShA9 + 100 CFU/cm2 on the non-lesional ventral arm skin of AD participants 
positive for S. aureus (AD SA+)  
2. The count of serious and non-serious treatment-emergent AEs per participant during the time period of Day  
0 to Day 31  
13.2.3 Exploratory Endpoints   
1. Association of S. hominis A9 bacteria abundance estimated CFU as measured by % S. hominis A9 positive 
colonies by PCR x CoNS CFU while CoNS CFU is above baseline density measured before application of ShA9 
+ 100 CFU/cm2 on the lesional and non-lesional, separately, ventral arm skin of AD participants between the 
arm treated with S. hominis A9 and the contralateral arm treated with placebo, and independently, with the 
participant’s face  
2. The duration of survival, measured as the time needed for CoNS CFU to drop below baseline density 
measured before application of ShA9 + 100 CFU/cm2 on the lesional ventral arm skin of AD participants 
negative for S. aureus (AD SA-)  
3. The duration of survival, measured as the time needed for CoNS CFU to drop below baseline density 
measured before application of ShA9 + 100 CFU/cm2 on the non-lesional ventral arm skin of AD participants 
negative for S. aureus (AD SA-)  
4. Comparison of time needed for CoNS CFU to drop below baseline density measured before application of 
ShA9 + 100 CFU/cm2 on the lesional and non-lesional, separately, ventral arm skin between AD participants 
positive (AD SA+) vs negative (AD SA-) for S. aureus   
5. Comparison of time needed for CoNS CFU to drop below baseline density measured before application of 
ShA9 + 100 CFU/cm2 on lesional vs non-lesional skin of AD participants positive (AD SA+) and negative (AD 
SA-) for S. aureus, separately  
6. Comparison of the L-EASI score of the ventral arms at Day 0 vs the last in-clinic visit among AD SA+, and 
independently, among AD SA-  
7. Comparison of the SCORAD score at Day 0 vs the last in-clinic visit among AD SA+, and independently, 
among AD SA-  
8. Comparison of the local Pruritus NRS of the ventral arms at Day 0 vs the last in-clinic visit among AD SA+, 
and independently, among AD SA-   


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9. Comparison of the proportion of antibiotic-sensitive CoNS to penicillin G, tetracycline, and erythromycin 
isolated from skin swabs at baseline and after ShA9 treatment, at the last swab collection from lesional skin  
  
13.3.  
Measures to Minimize Bias   
Bias will be minimized by stratifying by arm-dominance and then randomizing active-dominant, placebo-non-dominant 
vs placebo-dominant, active-non-dominant, where active is considered S. hominis A9.  
13.4.  
 Analysis Plan  
13.4.1 Analysis Populations  
Safety Sample and modified ITT sample will include all participants who are enrolled and receive any amount of 
S. hominis A9.  For analysis, S. aureus CFU results from swabs obtained prior to treatment application during the 
Treatment Visit will be used to define SA+ vs SA- status of participants.  
13.4.2 Primary Analysis of Primary Endpoint   
The primary endpoint for this pilot study is the time needed for CoNS CFU to drop below baseline density 
measured before application of ShA9 + 100 CFU/cm2 (time-to-CoNS elimination) in AD participants positive for S.  
aureus colonization.  Time-to-CoNS elimination is defined as the time from Day 0 to the first occurrence of a 
value below baseline density measured before application of ShA9 + 100 CFU/cm2.  The time to event for 
participants who did not reach CoNS CFU below baseline density measured before application of ShA9 + 100 
CFU/cm2 during the study will be right-censored.   
Time-to-CoNS elimination distribution (i.e. overall survival) will be estimated using Kaplan Meier techniques.  
Median survival time along with 95% CI will be constructed based on a log-log transformed CI for the survivor 
function S(t).  A graph of the Kaplan-Meier survival curve will be presented, and rates at fixed days will be 
derived (Therneau, et al., 2000).  Furthermore, estimation of the hazard rate and 95% confidence interval will be 
produced using a parametric survival model.  
  
13.4.3 Supportive Analyses of the Primary Endpoint   
A nonlinear time-response regression analysis will be fitted between CoNS CFU/cm2 and time, using either the 
two or three-parameter log-logistic model.  Each of the models will be fitted to the data, and their fit compared 
using the Akaike information criteria (AIC), where the model with the lowest AIC will be the primary model for 
the derivation of the ED50 (or T50, time at which half of the maximum CoNS CFU/cm2 is reached) (Ritz, et al., 
2015).  
13.4.4 Analyses of Secondary Endpoints  
Overall survival comparisons between culture-positive and culture-negative for S. aureus colonization will be 
performed using a two-sided log-rank test with a significance level of 10%.  Additionally, if the log-rank test is 
significant, the hazard ratio (HR) along with the 95% CI will be reported from a Cox proportional hazards 
regression model (discrete/exact method).  
The summary of all treatment-emergent adverse events and treatment-emergent serious adverse events will be 
provided by system organ class, high-level term, and preferred term in descending order of frequency, and by 
preferred term in descending order of frequency.  All analyses for adverse events will be conducted for the 
safety population by S. hominis A9 and Placebo.   


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13.4.5 Analyses of Exploratory Endpoints  
Repeated measures correlation will be calculated for determining the common within-individual association for 
paired (S. hominis treated arm and contralateral arm treated with placebo, and independently, with the 
participant’s face) for S. hominis A9 bacteria abundance estimated CFU as measured by % S. hominis A9 positive 
colonies by PCR x CoNS CFU assessed on two or more days.  The correlation coefficient, the 95% confidence 
interval, and the p-value will be reported.  Log transformation will likely be needed for the outcome of interest 
to produce a less skewed distribution (Bland and Altman, 1995).   
Analyses of disease severity (L-EASI, SCORAD, and local Pruritus NRS) change between Day 0 vs the last in-clinic 
visit among AD SA+, and independently, among AD SA- will be conducted using a repeated measures 
mixedeffects model with an unstructured covariance matrix.  The model will estimate the pre- vs. post-
intervention least squares means ± standard error and their difference reported with a 95% confidence interval 
and p-value.  
Analyses to investigate if ShA9 application can change CoNS antibiotic sensitivity to penicillin G, tetracycline, and 
erythromycin will be conducted on all participants (AD SA+ and AD SA-) by comparing the proportion of 
antibiotic-sensitive CoNS [(CFU without antibiotic containing plate - CFU with antibiotic containing plate) / CFU 
without antibiotic containing plate] before and after treatment. In particular, we will model the difference in 
proportions of antibiotic-sensitive CoNS between swabs taken at baseline and swabs taken at the last Follow-Up  
Visit.  
13.4.6 Descriptive Analyses   
Patient baseline characteristics will be tabulated.  Continuous measures will be reported using the mean, 
standard deviation, minimum, maximum, median, first and third quartiles, and sample size.  Categorical baseline 
and demographic characteristics and study disposition will be reported as frequencies and proportions.   
13.5.  
Interim Analyses   
13.5.1 Interim Analysis of Primary Endpoint  
The primary analysis, that is, the time needed for CoNS CFU to drop below baseline density measured before 
application of ShA9 + 100 CFU/cm2 (time-to-CoNS elimination) in AD participants positive for S. aureus 
colonization, will be generated when complete data for 50% of participants becomes available.  The purpose of 
this interim analysis is to inform the development of upcoming trials, especially with regards to their design and 
power/sample size considerations.  
13.5.2 Interim Analysis of Safety Data  
The DSMB will receive periodic safety reports on enrolled participants.  However, no formal interim analysis of 
safety data will be conducted.  The DSMB may request modifications to the protocol based on its review of the 
findings.  
13.5.3 Futility Analysis  
No formal interim analyses are planned for futility (Not Applicable).  
13.6.  
Statistical Hypotheses   
The main goal of this pilot study is the estimation of the time needed for the CoNS CFU to drop below baseline density 
measured before application of ShA9 + 100 CFU/cm2 (time-to-CoNS elimination).  There will be no formal statistical 
hypothesis test for the primary analysis of the primary endpoint.  


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13.7.  
Sample Size Considerations   
The proposed sample size for this study is 20 participants with 13 S. aureus positive and 7 S. aureus negative.  The 
sample size was derived from ADRN-08: Targeted Microbiome Transplant in Atopic Dermatitis where, using a parametric 
survival model with exponential distribution, we estimated a hazard rate (λ) of approximately 0.09.   This estimate was 
derived from a follow-up time of 11 days (shorter study) with 35 participants where nine (25%) were right-censored at 
trial end.  The goal for this trial is to estimate the hazard rate and its 95% confidence interval with precision for the width 
of the confidence interval as 0.20.  The 13 participants (9 events) proposed for the primary analysis were derived using 
these previous data as a starting point but considering more conservative estimates with a hazard rate of 0.15 and 30% 
right censoring, which produces a two-sided 95% confidence interval width equal to 0.194.  
14. Identification and Access to Source Data  
14.1.  
Source Data  
Source documents and source data are considered to be the original documentation where participant information, visit 
consultations, examinations, and other information are recorded.   Documentation of source data is necessary for the 
reconstruction, evaluation and validation of clinical findings, observations, and other activities during a clinical trial.   
14.2.  
Access to Source Data  
The site investigators and site staff will make all source data available to the DAIT/NIAID, authorized representatives of 
DAIT/NIAID, as well as to relevant health authorities.  Authorized representatives as noted above are bound to maintain 
the strict confidentiality of medical and research information that may be linked to identify individuals.  
15. Protocol Deviations  
15.1.  
Protocol Deviation Definitions  
Protocol Deviation – The investigators and site staff will conduct the study in accordance to the protocol; no deviations 
from the protocol are permitted.  Any change, divergence, or departure from the study design or procedures constitutes 
a protocol deviation.  As a result of any deviation, corrective and preventive actions will be developed by the site and 
implemented promptly.  
Major Protocol Deviation (Protocol Violation) - A Protocol Violation is a deviation from the IRB approved protocol that 
may affect the participant’s rights, safety, or well-being and/or the completeness, accuracy, and reliability of the study 
data.  In addition, protocol violations include willful or knowing breaches of human subject protection regulations or 
policies; any action that is inconsistent with the NIH Human Research Protection Program’s research, medical, and 
ethical principles; and a serious or continuing noncompliance with federal, state, local or institutional human subject 
protection regulations, policies, or procedures.  Examples of Major Protocol Deviations are described in the MOP.  
Non-Major Protocol Deviation - A non-major protocol deviation is any change, divergence, or departure from the study 
design or procedures of a research protocol that does not have a major impact on the participant's rights, safety, or well-
being, or the completeness, accuracy, and reliability of the study data.  
15.2.  
Reporting and Managing Protocol Deviations  
The study site principal investigator has the responsibility to identify, document, and report protocol deviations as 
directed by DAIT/NIAID.  However, protocol deviations may also be identified during site monitoring visits or during 
other forms of study conduct review.   


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Upon determination that a protocol deviation has occurred, the study staff will a) notify the site PI, b) notify the SACCC 
and c) will complete a Protocol Deviation form.  The protocol deviation form will document at minimum the date the 
deviation occurred, the date it was identified, a description of the event, whether the deviation resulted in an SAE/AE, PI 
signature, IRB report requirement, and documentation of a corrective action plan.  DAIT/NIAID may request discussion 
with the site PI to determine the effect of the protocol deviation on the study participant and his/her further study 
participation, the effect of the protocol deviation on the overall study, and corrective actions.  The PI will sign the paper 
source Protocol Deviation CRF, electronically sign Major Deviations in the electronic data capture (EDC) system, and 
submit the deviation to the local IRB per IRB regulations.  Major protocol deviations will be reported to the DSMB by the 
DAIT/NIAID Medical Monitor at the medical monitor’s discretion.   
16. Ethical Considerations and Compliance with Good Clinical Practice  
16.1.  
Quality Control and Quality Assurance  
The sponsor will review site processes for quality management of the study prior to enrollment, to include processes for 
data and biological specimen collection.  Expectations will be communicated to the site regarding study conduct.  In 
addition, all study staff are required to have GCP, Human Subject Protection, and ICH training.    
The investigator is expected to perform internal quality management of study conduct and documentation and is 
required to keep accurate records to ensure that the conduct of the study is fully documented and to ensure that all 
CRFs are completed for every participant entered in the trial.  
Plans for quality control of electronic data capture and data management will be detailed in the data management and 
data validation plans and will be reviewed by the sponsor prior to study onset.  The CRFs will be completed online via a 
web-based EDC system that has been validated and is compliant with Part 11 Title 21 of the Code of Federal Regulations 
(CFR).  Study staff at the site will enter information into the eCRFs, and the data will be stored remotely at a central 
database.  Quality control (QC) procedures will be implemented by the EDC system’s continuous monitoring of data and 
real-time detection and correction of errors.  All elements of data entry (i.e., time, date, verbatim text, and the name of 
the person performing the data entry) will be recorded in an electronic audit trail to allow all changes in the database to 
be monitored and maintained in accordance with federal regulations.  Any missing data or data anomalies will be 
communicated to the site(s) for clarification/resolution.   
The Sponsor will develop a risk-based monitoring plan to direct study monitoring.  Sponsor monitors will follow written 
Standard Operating Procedures (SOPs) to verify that the clinical trial is conducted, data are generated, and biological 
specimens are collected, documented (recorded), and reported in compliance with the protocol, International 
Conference on Harmonization Good Clinical Practice (ICH E6(R2)), and applicable regulatory requirements. The 
investigational site will provide direct access to all trial related sites, source data/documents, and reports for the 
purpose of monitoring and auditing by the sponsor, and inspection by local and regulatory authorities.  
16.2.  
Statement of Compliance  
This clinical study will be conducted using good clinical practice (GCP), as delineated in Guidance for Industry: E6 Good 
Clinical Practice Consolidated Guidance, and according to the criteria specified in this study protocol.  Before study 
initiation, the protocol and the informed consent documents will be reviewed and approved by the local IRB.  Any 
amendments to the protocol or to the consent materials will also be approved by the local IRB before they are 
implemented.  


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16.3.  
Informed Consent Process  
The consent process will provide information about the study to a prospective participant and will allow adequate time 
for review and discussion prior to his/her decision.  The principal investigator listed on the FDA 1572 or designee will 
review the consent and answer questions.  The prospective participant will be told that being in the trial is voluntary and 
that he or she may withdraw from the study at any time, for any reason.  The participant (or their legally acceptable 
representative) will read, sign, and date a consent form before undergoing any study procedures.  Consent materials will 
be presented in the participant’s primary language.  A copy of the signed informed consent form will be given to the 
participant.  
The consent process will be ongoing.  The consent form will be revised when important new safety information is 
available, the protocol is amended, and/or new information becomes available that may affect participation in the 
study.   
16.4.  
Privacy and Confidentiality  
Following the Health Insurance Portability and Accountability Act (HIPAA) guidelines, a participant’s privacy and 
confidentiality will be maintained throughout the study.  Each participant will be assigned a unique identification 
number, and these numbers rather than names will be used to collect, store, and report participant information.  All 
biological samples will be labeled with a unique identification number.  Data reported in medical journals or scientific 
meetings will be presented in aggregate for participants as a whole.  No individual participant will be identified in any 
way.  Site personnel will not transmit documents containing personal health identifiers (PHI) to the study sponsor or 
their representatives.  
17. Publication Policy  
The ADRN Publications Policy will apply to presentations and publications of the results of this trial  


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Appendix A: ADRN Standard Diagnostic Criteria (Version 3.0 09May2014)  
The following definitions will be used consistently throughout ADRN protocols. In children less than 4 years of age, the 
disease must be present for at least six months to minimize the likelihood of recruiting children with other eczematous 
disorders that mimic atopic dermatitis.  
I. Atopic Dermatitis (AD)1  A. Active Atopic Dermatitis (AD)1   
Participants must have Atopic Dermatitis (as defined below) within the last 3 months.  
B. Inactive Atopic Dermatitis (AD)1   
Participants must have an absence of Atopic Dermatitis (as defined below) within the last 12 months.   
C. Definition  
Participants must have, according to medical records, or based on a careful and credible history (provided by the 
participant, caregiver, parent, or guardian) or by physical exam by an ADRN investigator:  
1. Pruritus  
2. Eczema (acute, subacute, chronic)  


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a. Typical morphology and age-specific patterns which include:  
• 
Facial, neck, and extensor involvement in infants and children  
• 
Current or prior flexural lesions in any age group  
• 
Sparing groin and axillary regions   
b. Chronic or relapsing history  
c. Most participants will have the following clinical associations that add support to the diagnosis: • 
 Early age at onset  
• 
Atopy  
a. Personal and/or family history  
b. Immunoglobulin (IgE) reactivity  
• 
Xerosis  
Participants may have the following clinical associations which help to suggest the diagnosis of AD but are too 
nonspecific for defining or detecting AD for research or epidemiological studies:  
1. Atypical vascular responses (e.g., facial pallor, white dermographism, delayed blanch response) 2. 
Keratosis pilaris/hyperlinear palms/ichthyosis  
3. Ocular/peri-orbital changes  
4. Other regional findings (e.g., peri-oral changes/peri-auricular lesions) 5. Peri-follicular 
accentuation/lichenification/prurigo lesions  
  
II.  
References   
Eichenfield F, Hanifin J, Luger T, Stevens S, Pride H. Consensus Conference on Pediatric Atopic Dermatitis. J Am Acad 
Dermatology 2003;49:1088-95.   
  
   




 
 
2. 
Assessment of full inclusion and exclusion criteria will occur during the Screening Visit, after participants have consented to study participation.  
3. 
Participants who meet full inclusion and exclusion criteria including medication and therapy washouts, may complete the Screening and Pre-treatment Visits on the same 
day. The following assessments will not be repeated, if the Pre-treatment Visit is conducted on the same day as Screening: medical history, physical exam, AD severity 
assessment, concomitant medications, pregnancy test, and vitals.    
4. 
Upon completion of the Day 3 Follow-Up Visit, a participant will be notified prior to their next scheduled follow-up visit if they need to return to the clinic. A participant will 
not be required to complete the remaining follow up clinic visits (Day 10 through Day 24) if the lesional swabs from both arms (right and left) collected at their most recent 
prior visit, have CoNS CFU below baseline density measured before application of ShA9 + 100 CFU/cm2.    
 
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5. 
If disease activity increases, participants experience signs and symptoms as described on the instructional handcard, or other concerns arise between regularly scheduled 
visits, participants may be asked to return to the study site for an Unscheduled Visit.  
6. 
An abbreviated Medical History will be collected.   
7. 
AD disease severity will be assessed using the Investigator Global Assessment (Screening only), local Eczema Area and Severity Index (L-EASI) on the arms, SCORing Atopic 
Dermatitis (SCORAD), and local Pruritus Numerical Rating Scale (NRS).  
8. 
A urine pregnancy test will be completed for all female participants of child bearing potential who do not self-report as pregnant at the Screening and Treatment Visits.  
9. 
Vital signs will include temperature, heart rate, respiration rate, systolic blood pressure, and diastolic blood pressure. Growth parameters (height and weight) will only be 
collected at the Screening Visit.   
10. Skin swab collection from the participant’s ventral arms will occur during each visit, and skin swab collection from the participant’s face will begin at the Treatment Visit.  
11. Lesional and non-lesional sites will be identified on the participant’s ventral arms for skin swab collection. Digital photographs of the lesional and non-lesional swab sites 
will be taken prior to swab collection. Each photograph will include a ruler so the scale of the sites can be determined. These sites will be swabbed at subsequent visits.  
12. A non-lesional site will be identified on the participant’s face for skin swab collection. A digital photograph of the non-lesional swab site will be taken prior to swab 
collection. The photograph will include a ruler so the scale of the site can be determined. This site will be swabbed at subsequent visits.  
13. Participants will remain in clinic for up to 6 hours following the application of investigational product during the Treatment Visit. Additional skin swabs will be collected from 
the ventral arms at 15 minutes, 1, 2, 4, and 6 hours post application.   
14. Skin swabs and/or blood may be collected during an Unscheduled Visit per investigator discretion.   


 
 
 
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