Clinical trial NCT02075203
Conditions: Tuberculosis
Keywords: BCG Vaccinated
Sponsor: Aeras
Start date: 2014-02-01; Completion date: 2017-10-06
Enrollment: 989 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Randomized, Placebo Controlled, Partially Blinded Phase II Study to Evaluate Safety, Immunogenicity, and Prevention of Infection with Mycobacterium tuberculosis of AERAS-404 and BCG Revaccination in Healthy Adolescents

Detailed description:
This Phase II, randomized, 3-arm, placebo controlled, partially blinded, clinical trial will be conducted in 990 healthy, HIV-uninfected, QFT-GIT negative, previously BCG vaccinated adolescents. The trial will be conducted at the South African Tuberculosis Vaccine Initiative (SATVI) site in the Western Cape region of South Africa, where epidemiological studies involving thousands of adolescents have been conducted over the last decade to characterize rates of Mtb infection and active TB disease in this age group. Subjects will be enrolled in two sequential cohorts and within each cohort subjects will be randomized in a 1:1:1 ratio to receive either AERAS-404 or saline placebo on Days 0 and 56, or BCG Vaccine SSI on Day 0. The first 90 subjects (30 from each arm) will form the Safety \& Immunogenicity Cohort and will be subject to more intensive collection of safety data, with data reviewed by the Data Monitoring Committee (DMC), principal investigator and local medical monitor. Selected immunogenicity assays, including whole blood intracellular cytokine staining (ICS), will also be performed in this cohort. The remaining 900 subjects will be enrolled into the Correlates Cohort. All 990 subjects in the study will be evaluated for safety and biomarker outcomes, and for prevention of Mtb infection.

The primary Mtb infection endpoint will be QFT-GIT conversion from a negative to positive test, using the manufacturer's recommended threshold of 0.35 IU/mL, at any time-point after Day 84 and through end of follow-up for the primary endpoint. The 84-day 'wash-out' period is stipulated in order to exclude subjects who may have already been Mtb infected, but not yet converted their QFT-GIT test at screening, thus subjects who convert their QFT-GIT at Day 84 will not be included in the analyses of prevention of Mtb infection.

Arms: arm 1: 2 doses on Study Days 0 and 56 arm 2: 1 Dose on Study Day 0 arm 3: 2 Doses on Study Days 0 and 56
Interventions: intervention 1: AERAS-404 intervention 2: Placebo intervention 3: Bacillus Calmette-Guérin (BCG)
Intervention descriptions: intervention 1: The H4 antigen is a fusion protein created from two Mtb antigens: antigen Ag85B and TB10.4. Ag85B is also referred to as α-antigen and is a 30-kDa mycolyl transferase protein. TB10.4 is one of three members of the very similar ESAT-6 group of proteins found in Mtb culture supernatants. TB10.4 induces broad immune responses in T cells isolated from TB subjects compared to BCG-vaccinated donors and unvaccinated donors. IC31 is a combination of a leucine-rich peptide named KLK \& a synthetic oligonucleotide named ODN1a. The optimal molar ratio of KLK to ODN1a in mice is 25:1. AERAS-404 \& saline placebo trial arms will be double-blinded since BCG causes a recognizable local injection site reaction, the BCG revaccination trial arm will be unblinded. intervention 2: Saline intervention 3: BCG SSI Vaccine is registered in South Africa for prevention of TB in children and adults. BCG, an attenuated, live culture of the Bacillus Calmette-Guérin, was originally attenuated between 1906 and 1919 by serial passage of an M. bovis strain. The manufacturer Statens Serum Institut (SSI) in Copenhagen, Denmark derives this vaccine from the Danish BCG strain 1331. BCG SSI is supplied by the manufacturer in amber 10-dose vials containing 0.75 mg lyophilized SSI BCG. The BCG revaccination trial arm will be unblinded (open label).

Protocol documents:
ICF_002.pdf: 
 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 1 of 15 
                                                                                                                                                                 
PARTICIPANT INFORMATION AND CONSENT FORM 
 
Protocol Title:  
A Randomized, Placebo Controlled, Partially Blinded Phase 
II Study to Evaluate Safety, Immunogenicity, and 
Prevention of Infection with Mycobacterium tuberculosis 
of AERAS-404 and BCG Revaccination in Healthy 
Adolescents 
 
Protocol Number:  
C-040-404 
 
Short Title:  
Prevention of Infection with Mycobacterium tuberculosis 
of AERAS-404 and BCG Revaccination 
 
Study Doctor and Telephone:  
Dr Mark Hatherill (Principal Investigator) 
 
 
 
 
 
+27 21 404 7618 
 
Research Site:  
 
 
South African Tuberculosis Vaccine Initiative (SATVI) 
Brewelskloof Hospital, Haarlem Street, Worcester 6850 
South Africa 
 
 
Sponsor:  
 
 
 
Aeras, Rockville, Maryland, United States 
 
 
Introduction 
You have been asked to join a research study of an experimental vaccine that may help prevent 
tuberculosis (TB). TB is an infection caused by a bacteria (germ) called Mycobacterium 
tuberculosis.  TB is very common in South Africa. TB can spread from one person to another and 
can often cause death.  A vaccine is different than medicine.  It can prevent you from getting TB 
by helping your body fight the bacteria. 
 
You probably received the BCG vaccine against TB as a baby. Previous studies have shown that 
the BCG vaccine works in babies, but does not work very well to prevent TB in teenagers or 
adults. Vaccine researchers are looking for better vaccines, or better ways to use the BCG 
vaccine. The vaccine used in this study (AERAS 404) is called “experimental” because it is still 
being tested and we do not know if it works.  AERAS 404 vaccine is not approved for use or sold 
anywhere in the world. In this study we are looking at either vaccinating you with AERAS 404 or 
revaccinating you with the BCG vaccine. Some study participants will not get either the Aeras 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 2 of 15 
                                                                                                                                                                 
404 vaccine or the BCG, but will get a dummy vaccine called a “placebo”. The placebo is a 
harmless injection of salt water. Some participants will get the placebo so that we can compare 
their reactions to participants who did recieve one of the other 2 active vaccines.  We are 
researching if AERAS 404 (experimental vaccine) or revaccination with BCG vaccine can prevent 
teenagers from getting infected with the TB germ.  
 
Although all people who get sick from TB have been infected with the TB germ, only a few of  
the people who have been infected go on to become sick with TB. For this reason, and because 
infection with the TB germ is so common in South Africa, people who have been infected are  
only treated if they become sick with TB (unless they have an extra risk for TB, such as people  
with HIV infection and young babies). 
 
It is your choice whether or not to join the study. You do not have to be in the study if you do 
not want to. Consent means agreeing to take part in a research study.   
 
This consent form will tell you the purpose of this study, what you will be asked to do if you are 
in the study, and any possible harm to you from being in the study.  
 
Please ask the study doctor or a member of the study team to explain any words or other things 
you do not understand about the study.  Do not sign this consent form until you have answers 
to all of your questions.  You will be given a signed copy of this consent form. 
 
A summary of this study (in English) is on the website http:/www.ClinicalTrials.gov or  
http:/www.sanctr.gov.za.   This website does not include information that could identify 
anyone participating in this study.   You can search this website at any time.     
 
Why is this study being done? 
The main purpose of this study is to see if AERAS 404 (experimental vaccine) or revaccination 
with BCG vaccine can prevent adolescents from getting infected with the TB germ.  Some 
people are infected with TB and do not get sick or show symptoms.  We will test your blood to 
see if you are already infected with TB before you are enrolled in the study.  We want to study 
if either vaccine can prevent infection in participants who are not already infected with the TB 
germ.  
 
 
 
 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 3 of 15 
                                                                                                                                                                 
What are the experimental vaccines being tested in this study? 
The study vaccine is called AERAS-404.  AERAS-404 may prevent your body from getting TB.  
There is no danger of getting TB from an injection of AERAS-404.  The other vaccine is BCG.  It is 
approved to be used in babies, but in this study revaccination with BCG will be experimental. 
 
How many participants will be in this study?  How long will I be in this study? 
About 1000 participants in one country will be in this study.  You will be in the study for up to 
30 months (2 ½ years), depending on the results of your blood tests during the study. 
 
If I decide to join the study what happens first? 
If you want to join this study and you sign this consent form, you will be asked some questions 
and some tests will be done to see if you can be in the study.   
This will be done during a Screening Visit at the clinic.   
The following things will happen during the Screening Visit: 
 
• You will be asked to give the study staff a telephone number or a method to contact you 
during the study.  
• You will be asked if you are willing to complete all scheduled study visits. 
• You will be asked questions about your health and the medicines you take.   
• Your height, weight, temperature, and pulse rate will be measured. 
• You will have a physical exam. 
• The study staff may contact your regular doctor (if you have one) to get your medical 
history, and to inform the doctor about this study. 
• Females who are able to get pregnant will have a urine pregnancy test.   
• Your urine will be tested for routine lab tests.  
• You will be asked if you have /had TB or have been close to someone who has/had TB.  
If you have/had TB or have been exposed to someone who has TB, you cannot be in the 
study.  
• Blood will be collected for a TB test. This test will tell us if you are infected with the TB 
germ.  The TB test is not to see if you are sick and  have symptoms of TB.  
If this test is positive, you cannot be in this study.  
• The study staff will ask your permission to review your medical records and to check 
your HIV status.  If you have  ever had a positive HIV test you cannot be in this study.     
• Blood will be collected from you for an HIV test.  We will counsel you privately about 
HIV testing before the HIV test and we will counsel you about the meaning of the HIV 
test result afterwards. The results of the HIV test will be provided and explained to you 
after the counselling session in a confidential setting at the research clinic. If the result is 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 4 of 15 
                                                                                                                                                                 
positive, you cannot be enrolled into the study, but we will refer you to your local clinic 
for HIV treatment. If the HIV test is negative you can be in the study, depending on the 
results of the other tests. 
• The total amount of blood collected during these visits will be about 16 mls (1 
tablespoon). 
 
After the study staff reviews all this information they will decide if you can be in this study.  If 
you cannot be in the study, the study doctor will tell you why.   
 
How will this study be done? 
If you are eligible to be in this study you will receive either BCG, or the study vaccine AERAS 
404, or a placebo.  A placebo looks like the vaccine but does not have any vaccine in it.  Which 
group you will be in is decided by chance, like flipping a coin.  You have a 1 in 3 chance of 
getting BCG, the AERAS 404 study vaccine, or placebo.   
 
You will know if you are getting the BCG vaccine, but if it is decided by chance that you will get 
AERAS 404 or placebo, you will not know which one you receive. The study is done this way to 
compare the AERAS 404 vaccine to the placebo so that we know if any effects are caused by the 
vaccine.  
If you receive BCG, you will receive one injection (with a needle) in the upper arm. If you 
receive AERAS 404, or placebo, you will receive an injection in the upper arm on 2 different 
days (Study Visits) about 56 days apart.   
 
How long you stay in the study will depend on the results of  your blood tests. You will have to 
visit the clinic/hospital up to 15 different days for tests and evaluations no matter which 
vaccine you receive. 
 
What will happen at the study visits? 
You will have to visit the clinic up to 15 times during the study.  
Each of these visits is called a Study Visit.  
 
Blood will be collected from you on some of these Study Visits.  This 
photograph shows you how much blood is in “1 tube” of blood. One 
tube collects about 10 mLs of blood.   
 
At some visits one or more tubes of blood will be collected.  Three 
months after the first vaccination, we will start testing your blood 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 5 of 15 
                                                                                                                                                                 
to see if  you have been infected with the TB germ. If the test at three months shows that you  
have been infected you will only need to come back for one more Study Visit. If your blood test 
shows that you have been infected with the TB germ after the three months Study Visit (at 
Months 6 to 24), we will ask you to come back to the clinic two more times over the next six 
months to recheck your blood.  So you may participate up to 30 months. If you stay in the study 
for the whole 30 months, the total amount of blood collected from you during this study will be 
approximately 350 mls (23 tablespoons or 35 tubes).  
 
If you start to show symptoms of TB at any time during the study, we will test you for TB.  If the 
TB test results confirm that you have TB, the study doctor will advise you about your future 
medical care and treatment for TB disease. 
 
The table’s below shows what will happen at each Study Visit. 
 
Study Group BCG Revaccination 
 
Tests done on Study Days 0 to Month 24  
Study Visit 
Injection 
Blood 
Collected* 
Urine 
Collected 
Blood pressure, 
temperature, 
pulse rate 
taken 
Review 
Diary 
Card 
 
TB 
symptom 
screen 
 
HIV 
Test 
Screen 
 
16mls 
Yes 
Yes 
 
Yes 
Yes 
Day 0 
Yes 
53mls 
Yes 
Yes 
 
 
 
Day 7 
 
16mls 
Yes 
 
Yes 
 
 
Day 28 
 
 
 
 
 
Yes 
 
Day 70 
 
32.5mls 
 
 
 
Yes 
 
Day 84 
 
10mls 
 
 
 
Yes 
 
Month 6 
 
52.5mls 
 
 
 
Yes 
 
Month 12 
 
52.5mls 
 
 
 
Yes 
 
Month 18 
 
52.5mls 
 
 
 
Yes 
 
Month 24 
 
52.5mls 
 
 
 
Yes 
 
*All blood volumes are approximate 
 
 
 
 
 
 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 6 of 15 
                                                                                                                                                                 
Study Groups Aeras-404 and placebo 
 
Tests done on Study Days 0 to Month 24 
Study Visit Injection 
Blood 
Collected* 
Urine 
Collected 
Blood pressure, 
temperature, 
pulse rate taken 
Review 
Diary 
Card 
TB 
symptom 
screen 
HIV 
Test 
Screen 
 
16mls 
Yes 
Yes 
 
Yes 
Yes 
Day 0 
Yes 
53mls 
Yes 
Yes 
 
 
 
Day 3 
 
6mls 
Yes 
 
 
 
 
Day 7 
 
10mls 
Yes 
 
Yes 
 
 
Day 28 
 
 
 
 
 
Yes 
 
Day 56 
Yes 
 
Yes 
Yes 
Yes 
Yes 
 
Day 63 
 
10mls 
Yes 
 
Yes 
 
 
Day 70 
 
32.5mls 
 
 
 
 
 
Day 84 
 
10mls 
 
 
 
Yes 
 
Month 6 
 
52.5mls 
 
 
 
Yes 
 
Month 12 
 
52.5mls 
 
 
 
Yes 
 
Month 18 
 
52.5mls 
 
 
 
Yes 
 
Month 24 
 
52.5mls 
 
 
 
Yes 
 
*All blood volumes are approximate 
 
What else happens on the injection days? 
The following things will happen before the injection:  
• Females who are able to get pregnant will have another urine pregnancy test. If you are 
pregnant or nursing you cannot be in the study.    
• You will be asked if you have recently been vaccinated with any other vaccines. 
• Your blood pressure, temperature, and pulse rate will be measured.  
 
The following things will happen after the injection: 
• You must remain in the clinic for at least 30 minutes (half an hour) after receiving each 
injection to make sure you don’t have any bad side effects.  Your temperature and pulse 
rate will be taken and must be normal before you can leave the clinic. 
 
What are your duties as a study participant? 
Your duties as a study participant include the following: 
• You sign the informed consent if you understand it and want to remain in the study. 
(You previously signed an assent form when you were under 18). 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 7 of 15 
                                                                                                                                                                 
• Come to all Study Visits when you are told to.  You may have a total of up to 15 Study 
Visits.   
• Give complete and truthful information about your medical history and any signs of 
illness you may have during the study. 
• Give complete and truthful information about all medicines you take (or stop taking) 
during the study, including medicines from other doctors, herbal medicines, and 
medicine you buy yourself. 
• Give blood and urine samples to be tested for routine safety tests and responses to the 
vaccine.  
• Complete the daily diary and remember to bring it to the study clinic for review; 
• Tell the study doctor if you no longer want to be in the study. 
 
What are the possible harms?   
Expected side effects include those commonly seen with other vaccine injections, such as pain, 
redness, warmth, or swelling at the place where you get the injection.  These side effects 
usually last less than one week.  As with any new vaccine, unexpected and serious reactions, 
including allergic reactions to the vaccine, may occur. This could include an allergic reaction 
called anaphylaxis, which can be described as feeling dizzy, with difficulty in breathing, an itchy 
throat, or tingling arms. If you should experience any of these symptoms, you need to contact 
the study doctor or the nearest doctor immediately. This vaccine has been tested before in 
both humans and animals, and anaphylaxis did not occur in any of them. 
 
As a result of the BCG vaccine, you may also experience a small sore that might grow into a 
circle and may become crusty, and which can take up to three months to go away. A small scar 
may also develop at the site of vaccination. The following are some of the possible other side 
effects of vaccination: 
• Headache  
• Fever 
• Tiredness  
• Enlargement of glands  
Other less likely side effects from vaccines include diarrhea and nausea. Other side effects may 
also occur that are unknown at this time. 
 
Very rarely, people who receive BCG can have bone infections or severe local infections called 
abscesses.  These severe reactions usually happen in people who have other medical problems, 
like HIV infection. We do not expect to see these kinds of severe reactions in this study.   
 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 8 of 15 
                                                                                                                                                                 
When blood is collected you might feel brief pain or burning where the needle enters the skin.  
Your skin might be bruised and you may feel dizzy. You might faint, but this rarely happens. 
During the study you will receive general check-ups and a study team member will be available 
to evaluate any side effects and answer your questions.  We will teach you about the symptoms 
of TB, and will ask you if  you have any of these symptoms. If we think you might have TB we 
will help you get medical care.    
 
This is not the first study of Aeras-404 in humans. 196 persons in total have received Aeras-404.  
In other research studies with Aeras-404, some participants experienced redness at the place 
where the injection was given. Pain at the place where the injection was given is common. 
Aeras-404 has also been tested and shown to be safe in animals such as mice, rabbits, and 
guinea pigs.   
 
For females  
• Females who are pregnant or breastfeeding cannot be in the study because it is not 
known if the study vaccine is safe for pregnant women, fetuses, and breastfeeding 
children. You will not be given an injection if you have a positive pregnancy test at 
Screening or on an injection day.  
• If you are able to become pregnant, it is very important that you do not become 
pregnant during the study.  You must agree to the following acceptable methods of 
birth control starting 28 days prior to the first study vaccine injection and for the 24 
months period during the study.  
  
o Acceptable methods of avoiding pregnancy include a sterile sexual partner, 
sexual abstinence (not engaging in sexual intercourse), hormonal contraceptives 
(oral, injection, transdermal patch, or implant), vaginal ring, intrauterine device 
(IUD), or the combination of a condom or diaphragm with spermicide 
• If you think you have become pregnant while in the study, you must contact the study 
doctor immediately. The study doctor will advise you about your baby’s future medical 
care. 
 
What will happen to my samples? 
All the samples collected in this study (like blood or urine) will be stored up 5 years after the 
study is finished.  The samples will only be used for tests needed for this study.  You will be 
asked later if we can use your samples for other tests.  The samples will be stored at the SATVI 
laboratory at the University of Cape Town, at the Aeras South Africa Endpoint Assay lab in Cape 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 9 of 15 
                                                                                                                                                                 
Town, or the Aeras laboratory in the USA. We may transfer some of the stored samples to other 
places too to do the tests. 
 
Are there any benefits to me from being in the study? 
There are no medical benefits to you for being in this study except you may receive medical 
tests that you do not usually get. The information learned from this study may lead to the 
development of a new vaccine to prevent TB that could benefit society. 
 
Are there any alternative to being in this study? 
Your alternative is not to be in this study. 
 
How can I leave the study? 
Being in this study is your choice. You may choose not to be in the study.  You can also leave the 
study at any time.  If you decide to leave the study, you should tell the study doctor 
immediately. If you do not want to join the study, or wish to stop being in the study before it is 
over, it will not affect the medical care you have already received.  If you wish to leave the 
study early, we may ask you to come for one final study visit to do a final physical examination 
and to do some final blood tests. If new information about the study vaccine becomes available 
that might affect your decision to participate in the study, you will be told right away. 
 
Could I be taken out of the study?  Could the study be stopped? 
You may be taken out of the study if the study doctor decides this would be best for you or if 
you do not follow the study rules.  The study sponsor or a government health authority like the 
Medicines Control Council could also decide to stop the study at any time.   
 
Who is paying for this study? 
Aeras and Sanofi-Pasteur are paying for the study vaccine, placebo, tests, medical exams, and 
procedures in this study.  Your insurance company will not have to pay anything for you to be in 
this study.  The study doctor(s) and study staff do not have a direct financial interest in Aeras or 
the Aeras-404 vaccine. 
 
Will I be paid for being in the study? 
You will receive money as compensation for your time, inconvenience, and any additional costs 
for being in this study. You will receive R150 per visit during this study.  
 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 10 of 15 
                                                                                                                                                                 
Samples (like blood or urine) collected from you during this study may be used to develop 
products that could be patented or sold in the future.  You will not receive any money or other 
benefits from these products.   
 
What if I am injured because of this study?  
If you suffer a physical injury from this study, the South African TB Vaccine Initiative (SATVI) will 
give you immediate medical treatment.  SATVI may also refer you to a doctor or clinic for 
further care.   
 
Aeras has taken out insurance in the event of a study-related injury to you. This insurance 
follows the Association of the British Pharmaceutical Industry Guidelines. The guidelines 
recommend that the sponsor compensates you for any injury that is caused by the study 
vaccine or study procedures.  You do not have to prove that the company is at fault.  
 
Aeras will not pay to treat a medical condition or disease you had before joining this study or 
expenses for injury, treatment, or hospitalization you may require that are not the result of 
your participation in the study.   
 
No other compensation, such as lost wages or other damages, will be available.  You do not 
waive any of his/her legal rights by you signing this consent form. 
 
You may make a claim from the sponsor's insurer. If successful, you may be asked to say that 
you won't make further claims. But even if you say so at that time, you still have the right to 
make a separate claim against the sponsor in a South African court. 
 
How will my personal information and rights be protected? 
Your samples (like blood and urine) will be stored and tested for this study.  Your name will not 
appear on the samples. 
 
Your study records will contain personal health information.  Aeras (and their representatives) 
will visit the clinic/hospital from time to time to check that the study is being done correctly and 
will review your study records.  You will not be identified by name in these study records.  
However, Aeras has the right to see your name, if they need to do so.  Aeras will not share your 
study records or identity with other parties unless it is required by law.  Aeras will do everything 
they can to ensure that your study records and samples are protected.  However, we cannot 
promise that records and samples will be fully protected. 
 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 11 of 15 
                                                                                                                                                                 
Your study records may also be reviewed by the Human Research Ethics Committee, University 
of Cape Town.  The Ethics Committee will make every reasonable attempt to keep your identity 
confidential and will not share your study records with other parties unless it is required by law.  
 
Your study records and medical records may also be reviewed by representatives from 
government agencies or other health authorities of South Africa. Your study records will be 
kept at the clinic/hospital for an undetermined amount of time following the completion of the 
study.  After the study is completed (1) you may see your records, (2) you will be told whether 
you received the study vaccine or the placebo, and (3) you will be told the results of the study.  
If you decide to leave the study, information already collected from you may still be used for 
the study. 
 
Aeras, the study doctor, or other doctors involved in the study may publish reports or articles 
on the study or present the study findings to scientific groups.  Your study records may be 
provided to the people preparing the reports and may be included in the reports.  However, 
your identity and name will not be revealed in any of those reports, articles, or presentations. 
 
Who can I contact for information about the study? 
If you have questions about the study you can contact the Investigator on site, Dr Hennie 
Geldenhuys, or other study team members by calling 076 692 1837. 
 
If you have any symptoms or injuries you think might be related to being in this study contact 
Dr Hennie Geldenhuys the 24-hour telephone number to call is 076  692 1837 
 
If you have any additional questions about your rights as a research participant, you should 
contact the University of Cape Town Human Research Ethics Committee, which is overseeing 
the conduct of this study at The South African TB Vaccine Initiative.  An Ethics Committee is an 
independent committee established to help protect the rights of research participants 
If your doctor or the Ethics Committee did not provide you or you with answers to your 
satisfaction, you should write to the South African Medicines Control Council (MCC). 
 
 
Contact person 
The Ethics Committee Chairperson 
The Registrar 
Address 
University of Cape Town Faculty of Health 
Sciences, Human Research Ethics Committee 
Old Main Building, Groote Schuur Hospital, 
OBSERVATORY 7925 
Medicines Control Council 
Private Bag X828,  
Pretoria, 0001 
Telephone number 
021 406 6338 or 021 406 6626 
012 395 8000 
Fax number 
021 406 6411 
012 395 8468 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
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CONSENT TO PARTICIPATE IN A RESEARCH STUDY 
 
AERAS C-040-404, A Randomized, Placebo Controlled, Partially Blinded Phase II Study to 
Evaluate Safety, Immunogenicity, and Prevention of Infection with Mycobacterium 
tuberculosis of AERAS-404 and BCG Revaccination in Healthy Adolescents 
 
I have had the chance to read this consent form, discuss this study and ask questions.  My 
questions have been answered to my satisfaction.  I understand that by signing this consent 
form, I do not give up any of my legal rights.   
 
I hereby consent to be in this study.    I will receive a copy of this signed consent form.    
 
________________________________________________ 
Printed name of participant 
 
________________________________________________               ______________________ 
Signature/mark or thumbprint of participant 
Date (MM/DD/YY) & time 
 
_________________________________________________________________  
Printed name and designation of person conducting informed consent discussion  
 
 
______________________________________________ 
______________________ 
Signature of person conducting informed consent discussion 
 Date (MM/DD/YY) & time 
 
________________________________________________ 
*Printed name of impartial witness  
 
________________________________________________ 
______________________ 
*Signature of impartial witness                                                            Date (MM/DD/YY) & time 
 
*(Required only if participant or legally appointed representative is illiterate, an interpreter is used, or if 
required by another party.) 
  
 
This original signed consent form will remain in the files of the doctor conducting the study.   
A copy of the signed consent form will be given to the study participant. 
 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 13 of 15 
                                                                                                                                                                 
CONSENT TO USE SAMPLES FROM STUDY FOR FUTURE TB RESEARCH 
 
Sample Storage 
We are asking permission to store samples collected from you during this study (such as blood, 
urine, etc.) and use them for future TB research.  This research would not be part of this study.  
As new tests become available for TB research, your samples would be used to help learn more 
about TB and how to prevent it and treat it.   
 
Your decision whether or not to allow your samples to be stored and used for future TB 
research is separate from your choice to join this study.  If you choose NOT to allow your 
samples to be used for future research you can still join this study. 
 
Control of Samples and Privacy 
If you give permission, your samples will be placed in a secure storage facility.  Aeras  will be 
responsible for managing the facility.  Your name and any other information that could identify 
you will be removed from the samples before storage.  The sample will be identified by a 
number or barcode and the information linking you to this number/barcode will be held under 
safe, secure conditions by SATVI. If as a result of the future research, Aeras becomes aware of 
information which is clinically relevant and could affect the health and wellbeing of the 
participant they will supply the information to the doctor at SATVI who will then link the 
number / barcode to the participant and contact the participant for an appointment where this 
information can be discussed.   
 
Your samples will be stored for an undetermined amount of time. 
 
If you agree to have your samples stored and then later change your mind you can contact Dr 
Geldenhuys on 023 346 5400  to ask that your samples not be used and be destroyed.  The 
results of any tests done on your samples before the request is received will be kept by the 
researchers. 
 
Risks  
There is a risk that your samples (and any test results) could be linked to you.  This could impact 
your job, insurance, or family relationships.  The chance of this happening is very small. Aeras 
will do their best to protect your privacy and keep your identity secret. 
 
 
 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 14 of 15 
                                                                                                                                                                 
Benefits 
You will not receive any direct benefit by allowing your samples to be used for future TB 
research.  It may benefit others if knowledge is gained through the research. 
 
Tests done on your samples may help Aeras or other researchers develop a commercial 
product. You will not receive any financial profit if this occurs. Any publication resulting from 
this future research will not include any names or other identifiers of participants. 
 
Participant agreement: 
If any of the blood samples I have provided for this research project are unused or left over 
when the research is completed,  (tick ONE choice from each of the following boxes): 
□ I want my samples destroyed immediately 
OR 
□ I give permission for my samples to be stored indefinitely. 
AND  
if the sample is to be stored: 
□ I give permission for my samples to be stored and used in future TB research  
AND 
□ I am willing to be re-contacted by the researcher about possible future use of more blood 
samples in future research. 
OR 
□ I do not want to be re-contacted to ask me to give more blood samples in the future or to 
take part in future studies. 
 
 
 
 
 
 
 
 
 
 
 


 
 
 
 
 
 
Protocol C-040-404   
SATVI Participant ICF   
Version 4.0 
10  January 2014 
 
 
 
 
 
 
 
 
SATVI Site-specific Participant Information and Consent Form, Version 4.0 dated 10 January 2014 
Approved by the University of Cape Town Human Research Ethics Committee on 23 January 2014 
 
    Page 15 of 15 
                                                                                                                                                                 
I have read the information, or it has been read to me. I have been informed that my sample 
will be identified by a number / barcode and not my name. I have had the chance to ask 
questions about it and I am satisfied with the answers I was given. I consent voluntarily and 
understand that I have the right to withdraw my consent without this affecting the research I 
am currently taking part in or my medical care. 
 
________________________________________________ 
Printed name of participant 
 
________________________________________________               ______________________ 
Signature/mark or thumbprint of participant 
Date (MM/DD/YYYY) & time 
 
________________________________________________ 
 
Printed name and designation of staff member conducting informed consent discussion  
 
________________________________________________ 
______________________ 
Signature of person conducting informed consent discussion 
 Date (MM/DD/YYYY) & time 
 
________________________________________________ 
*Printed name of impartial witness  
 
________________________________________________ 
______________________ 
*Signature of impartial witness                                                                  Date (MM/DD/YYYY) & time 
 
*(Required only if participant or legally appointed representative is illiterate, an interpreter is used, or if   
required by another party.) 
  
 
 
 
 
 
 

Prot_000.pdf: 
 
 
Page 1 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
 
 
A Randomized, Placebo Controlled, Partially Blinded Phase II Study to Evaluate 
Safety, Immunogenicity, and Prevention of Infection with Mycobacterium 
tuberculosis of AERAS-404 and BCG Revaccination in Healthy Adolescents 
 
 
Investigational Product:  
 
AERAS-404 
Bacille Calmette Guerin (BCG) Vaccine SSI 
 
Aeras Protocol Number:  
 
C-040-404 
 
 
 
 
Sponsor: 
Aeras 
1405 Research Boulevard 
Rockville, Maryland 20850 USA 
Phone: 001-301-547-2900  
Fax: 
001-301-547-2901 
 
 
Sponsor's Authorized Representative: 
Ann M Ginsberg, MD, PhD 
Chief Medical Officer 
 
 
 
 
 
 
 
 
Principal Investigator Agreement: 
I, the undersigned, have reviewed this protocol and agree to conduct this protocol in accordance 
with Good Clinical Practices (ICH-GCP), the ethical principles set forth in the Declaration of 
Helsinki, and with local regulatory requirements.   
 
Signature 
 
 
 
 
 
 
 
Date 
 
 
Printed Name  


 
 
Page 2 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
 TABLE OF CONTENTS 
 
LIST OF ABBREVIATIONS ...................................................................................................... 5 
STUDY ABSTRACT .................................................................................................................... 7 
1 
INTRODUCTION........................................................................................................... 11 
1.1 
Background ............................................................................................................. 12 
1.2 
Description of AERAS-404 Vaccine ...................................................................... 13 
1.3 
Nonclinical Experience with AERAS-404 (HyVac4) Vaccine ............................... 14 
1.4 
Clinical Experience with AERAS-404 .................................................................... 14 
1.5 
Clinical Experience with Vaccines Related to AERAS-404 ................................... 17 
1.6 
Description of BCG ................................................................................................. 18 
1.7 
Rationale for Study .................................................................................................. 19 
2 
STUDY OBJECTIVES AND DESIGN ......................................................................... 20 
2.1 
Objectives ................................................................................................................ 20 
2.2 
Design and Endpoints .............................................................................................. 21 
3 
STUDY PROCEDURES ................................................................................................ 24 
3.1 
Schedule of Subject Evaluations ............................................................................. 24 
3.2 
Subject Selection ..................................................................................................... 27 
3.2.1 
Recruitment and Informed Consent .......................................................... 27 
3.2.2 
Screening................................................................................................... 28 
3.2.3 
Inclusion Criteria ...................................................................................... 28 
3.2.4 
Exclusion Criteria ..................................................................................... 29 
3.2.5 
Screening Clinical Assessments and Laboratory Tests ............................ 30 
3.3 
Study Randomization .............................................................................................. 30 
3.4 
Blinding ................................................................................................................... 31 
3.4.1 
Unblinding for Clinical Emergencies ....................................................... 31 
3.5 
Vaccine Administration ........................................................................................... 32 
3.6 
Study Evaluations .................................................................................................... 33 
3.6.1 
Evaluation of Mtb infection ...................................................................... 33 
3.6.2 
Immunology Laboratory Evaluations ....................................................... 34 
3.6.3 
Safety Evaluations .................................................................................... 36 
3.6.3.1 
Pre-vaccination and Post-Vaccination Monitoring of 
Subjects .................................................................................... 36 
3.6.3.2 
Clinical Assessments and Laboratory Tests ............................. 36 
3.6.3.3 
Adverse Events......................................................................... 37 
3.6.3.4 
Concomitant Medications ........................................................ 37 
3.6.4 
Subject Follow-up and Contact ................................................................. 38 
3.6.5 
Loss to Follow-up ..................................................................................... 38 
4 
VACCINES ...................................................................................................................... 38 
4.1 
Supplies ................................................................................................................... 38 


 
 
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Protocol C-040-404  
 
CONFIDENTIAL 
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Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
4.2 
Accountability ......................................................................................................... 39 
4.3 
Receipt and Storage ................................................................................................. 39 
4.4 
Vaccine Preparation ................................................................................................ 40 
4.5 
Disposal of Unused Supplies ................................................................................... 40 
5 
SAFETY ........................................................................................................................... 40 
5.1 
Responsibilities for Ensuring the Safety of Trial Subjects ..................................... 40 
5.1.1 
Principal Investigator ................................................................................ 40 
5.1.2 
Study Sponsor ........................................................................................... 41 
5.1.3 
Local Medical Monitor ............................................................................. 41 
5.1.4 
Data Monitoring Committee ..................................................................... 41 
5.1.5 
Institutional Review Boards and Ethics Committees................................ 41 
5.1.6 
National Regulatory Authority ................................................................. 42 
5.2 
Safety Surveillance During the Study ..................................................................... 42 
5.3 
Definition of Adverse Event ................................................................................... 42 
5.4 
Assessing Severity ................................................................................................... 43 
5.5 
Assessing Causal Relationship (Relatedness) ......................................................... 44 
5.6 
Definition of Adverse Reaction ............................................................................... 45 
5.7 
Solicited Adverse Events and Injection Site Reactions .......................................... 45 
5.8 
Adverse Events of Special Interest (AESIs) ........................................................... 46 
5.9 
Assessing "Seriousness" and Serious Adverse Events ............................................ 46 
5.10 Assessing Expectedness .......................................................................................... 47 
5.11 Definition of Suspected Unexpected Serious Adverse Reaction (SUSAR) ............ 47 
5.12 Reporting of Serious Adverse Events ..................................................................... 47 
5.13 Other Events Requiring Immediate Reporting ........................................................ 48 
5.14 Adverse Event Treatment, Follow-up, and Outcome .............................................. 49 
5.15 Follow-up of Subjects Who Become Pregnant ....................................................... 49 
5.16 Subject Diary and Daily Temperature Monitoring.................................................. 50 
6 
PAUSING AND STOPPING RULES ........................................................................... 51 
6.1 
Rules for Discontinuing Study Injections in an Individual Participant ................... 51 
6.2 
Rules for Suspension of the Entire Study ................................................................ 51 
7 
STATISTICAL CONSIDERATIONS .......................................................................... 52 
7.1 
Subject Populations ................................................................................................. 52 
7.2 
Demographics and Protocol Compliance ................................................................ 52 
7.3 
Prevention of Infection (QFT-GIT Conversion) Analyses...................................... 53 
7.4 
Immunogenicity and Other Immunology Analyses ................................................ 53 
7.4.1 
Immune Response Determined by Intracellular Cytokine 
Staining Assay .......................................................................................... 54 
7.4.2 
Additional Exploratory Analyses .............................................................. 54 
7.4.2.1 
RNA Transcriptomics and Whole Blood Absolute 
Blood Cell Counts .................................................................... 54 
7.4.2.2 
Quantification of Soluble Immune Mediators by 
Multiplex ELISA ...................................................................... 55 
7.4.2.3 
Correlates of Risk/Correlates of Protection ............................. 55 


 
 
Page 4 of 91 
 
 
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Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
7.4.2.4 
Novel IGRAs ............................................................................ 55 
7.5 
Safety Analyses ....................................................................................................... 55 
7.5.1 
Adverse Events ......................................................................................... 55 
7.5.2 
Clinical Laboratory and Vital Sign Parameters ........................................ 56 
7.6 
Sample Size Considerations .................................................................................... 56 
7.7 
Plan for Statistical Summaries and Analyses .......................................................... 57 
7.7.1 
Primary Data Review ................................................................................ 57 
7.7.2 
Final Study Report .................................................................................... 57 
7.8 
Computer Methods .................................................................................................. 57 
8 
DATA COLLECTION, MONITORING, AND RECORD RETENTION................ 58 
9 
HUMAN SUBJECTS ...................................................................................................... 58 
9.1 
Ethics and Regulatory Considerations .................................................................... 58 
9.2 
Institutional Review Board or Independent Ethics Committee ............................... 60 
9.3 
Informed Consent and Assent ................................................................................. 61 
10 
STUDY COMPLETION ................................................................................................ 62 
11 
PUBLICATIONS ............................................................................................................ 62 
12 
CHANGES IN THE PROTOCOL ................................................................................ 62 
12.1 Changes from Version 1.0 to Version 2.0 (Amendment 1)..................................... 62 
12.2 Changes from Version 2.0 to Version 3.0 (Amendment 2)..................................... 65 
12.3 Changes from Version 3.0 to Version 4.0 (Amendment 3)..................................... 67 
12.4 Changes from Version 4.0 to Version 5.0 (Amendment 4)..................................... 68 
13 
REFERENCES ................................................................................................................ 70 
APPENDIX A 
Detailed Description of Study Visits ...................................................... 74 
SAE and SUSAR Reporting Schemes ....................................................................................... 81 
APPENDIX B 
Toxicity Table (Modified from Division of AIDS Table for 
Grading the Severity of Adult and Pediatric Adverse 
Events) ...................................................................................................... 83 
APPENDIX C 
Adverse Events of Special Interest ........................................................ 90 
 


 
 
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Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
LIST OF ABBREVIATIONS 
 
βHCG 
beta human chorionic gonadotropin 
µg 
microgram(s) 
Ab 
antibody 
AE 
adverse event(s) 
AESI 
adverse event of special interest 
ALP 
alkaline phosphatase 
ALT 
alanine aminotransferase 
AST  
aspartate aminotransferase 
BCG 
Bacillus Calmette-Guérin 
BMI 
body mass index 
BUN 
blood urea nitrogen 
CBC 
complete blood count 
CFR 
Code of Federal Regulations (US) 
CRF 
case report form(s) 
DMC 
Data Monitoring Committee 
DOH 
Department of Health 
EDC 
electronic data capture 
ELISA 
enzyme-linked immunosorbent assay 
ELISPOT 
enzyme-linked immunospot 
GCP 
Good Clinical Practices 
HIV 
human immunodeficiency virus 
IB 
Investigator’s Brochure 
ICH 
International Conference on Harmonization of Technical 
Requirements for Registration of Pharmaceuticals for Human Use 
ICS 
intracellular cytokine staining 
ID 
intradermal 
IEC 
Independent Ethics Committee 
IFN-γ 
interferon gamma 
IGRA 
interferon gamma release assay 
IL-2 
interleukin-2 
IM 
intramuscular 
INH 
isoniazid 
IPT 
isoniazid preventive therapy 
IRB 
Institutional Review Board 
IUD 
intrauterine device 
LLN 
lower limit of normal 
MedDRA 
Medical Dictionary for Regulatory Activities 
mL 
milliliter(s) 
Mtb 
Mycobacterium tuberculosis  
NTP 
National TB Programme 
NTM 
non-tuberculous mycobacteria 
PBMC 
peripheral blood mononuclear cell(s) 


 
 
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CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
QFN 
QuantiFERON 
QFT-GIT 
QuantiFERON-TB Gold in-Tube 
RNA 
ribonucleic acid 
SAE 
serious adverse event(s) 
SAER 
supplemental serious adverse event report 
SATVI  
South African Tuberculosis Vaccine Initiative 
SOC 
system organ class 
SUSAR 
suspected unexpected serious adverse reaction(s) 
TB 
tuberculosis 
TNF 
tumor necrosis factor 
TST 
tuberculin skin test 
ULN 
upper limit of normal 
US 
United States 
WBC 
white blood cell 
WHO 
World Health Organization 
 


 
 
Page 7 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
STUDY ABSTRACT 
 
TITLE: 
A Randomized, Placebo Controlled, Partially Blinded Phase II Study to Evaluate Safety, 
Immunogenicity, and Prevention of Infection with Mycobacterium tuberculosis of AERAS-404 
and BCG Revaccination in Healthy Adolescents 
 
RATIONALE: 
This clinical trial will evaluate safety, immunogenicity, and prevention of Mtb infection (QFT-
GIT conversion), of AERAS-404 and BCG revaccination in previously BCG vaccinated 
adolescents. QFT-GIT negative adolescents in the area where this study will be conducted have 
previously been shown to have very high rates of TST and QFT-GIT conversion, associated with 
increased risk of active TB disease in subsequent years. A TB vaccination strategy incorporating 
BCG revaccination or AERAS-404 vaccination in adolescents or young adults, if found to 
prevent Mtb infection, would likely have a major impact on TB disease, TB transmission, and 
control of the epidemic. If revaccination with BCG or vaccination with AERAS-404 is shown to 
prevent infection with Mtb in this proof of concept study in adolescents, additional larger scale 
studies examining the impact on TB disease in more diverse populations would be warranted. 
 
OBJECTIVES: 
Primary Objectives:  
• To evaluate the safety profile in HIV-uninfected, remotely BCG vaccinated adolescents of 
o AERAS-404 
o BCG revaccination 
• To evaluate prevention of Mtb infection, as measured by rates of conversion using a QFT-
GIT assay, by 
o AERAS-404 compared to placebo 
o BCG revaccination compared to placebo 
 
Secondary Objectives 
• To evaluate prevention of Mtb infection, as measured by rates of sustained conversion using 
a QFT-GIT assay, by  
o AERAS-404 compared to placebo 
o BCG revaccination compared to placebo 
• To investigate the immunogenicity in HIV-uninfected, remotely BCG vaccinated adolescents 
of 
o AERAS-404 
o BCG revaccination 
 
DESIGN: 
This Phase II, randomized, 3-arm, placebo controlled, partially blinded, clinical trial will be 
conducted in 990 healthy, HIV-uninfected, QFT-GIT negative, previously BCG vaccinated 
adolescents. The trial will be conducted at the South African Tuberculosis Vaccine Initiative 
(SATVI) site in the Worcester region of the Western Cape region of South Africa, where 


 
 
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Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
epidemiological studies involving thousands of adolescents have been conducted over the last 
decade to characterize rates of Mtb infection and active TB disease in this age group. Additional 
sites with similar TB epidemiology may also be used. 
 
Subjects will be enrolled in two sequential cohorts and within each cohort subjects will be 
randomized in a 1:1:1 ratio to receive either AERAS-404 or saline placebo on Days 0 and 56, or 
BCG Vaccine SSI on Day 0. The first 90 subjects (30 from each arm) will form the Safety & 
Immunogenicity Cohort and will be subject to more intensive collection of safety data, with data 
reviewed by the Data Monitoring Committee (DMC), principal investigator and local medical 
monitor. Selected immunogenicity assays, including whole blood intracellular cytokine staining 
(ICS), will also be performed in this cohort. The remaining 900 subjects will be enrolled into the 
Correlates Cohort.  All 990 subjects in the study will be evaluated for safety and biomarker 
outcomes, and for prevention of Mtb infection. 
 
The primary Mtb infection endpoint will be QFT-GIT conversion from a negative to positive 
test, using the manufacturer’s recommended threshold of 0.35 IU/mL, at any time-point after 
Day 84 and through end of follow-up for the primary endpoint (see Figure 2-1). The 84-day 
‘wash-out’ period is stipulated in order to exclude subjects who may have already been Mtb 
infected, but not yet converted their QFT-GIT test at screening, thus subjects who convert their 
QFT-GIT at Day 84 will not be included in the analyses of prevention of Mtb infection.   
 
The duration of the trial will be endpoint-driven, using a design targeting accrual of 64 primary 
endpoints. The primary analysis will be triggered when at least 64 endpoints are accrued in the 
study (both cohorts) AND the median follow up time is at least 15 months, with a maximum 
individual follow-up time to detect initial QFT-GIT conversion of 24 months. If an initial 
reduction in infection rate in the BCG or AERAS-404 groups is seen in the primary analysis of 
QFT-GIT conversion, follow up for the entire study may be extended to examine the duration of 
prevention of Mtb infection. 
 
The trial sample size of 990 subjects is determined by the primary prevention of Mtb infection 
objective (measured by prevention of conversion to a positive QFT-GIT test). A potential 
mechanism of action of vaccination might be to allow initial Mtb infection (and QFT-GIT 
conversion), but to prevent establishment of sustained latent Mtb infection (and thus persistent 
QFT-GIT conversion), resulting in QFT-GIT reversion to negative. Thus the study will also 
examine the persistence of QFT-GIT conversion.  
 
All subjects with primary QFT-GIT conversion will be followed for an additional 6 months post-
conversion to ascertain the sustained QFT-GIT conversion and QFT-GIT reversion endpoints. 
The secondary Mtb infection endpoint will be the combination of QFT-GIT conversion from a 
negative to positive test, using the manufacturer’s recommended threshold of 0.35 IU/mL, at any 
time-point after Day 84 and through end of follow-up for the primary endpoint, AND persisting 
without QFT-GIT reversion from a positive to a negative test through 6 months after QFT-GIT 
conversion (see Figure 2-1).  
 


 
 
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Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
Immunological outcomes will include intracellular cytokine staining (ICS) on PBMCs and whole 
blood, transcriptomics on RNA from whole blood, absolute blood cell subset counting by flow 
cytometry (for deconvolution of transcriptomics), quantification of soluble immune mediators by 
multiplex ELISA (all in the Safety & Immunogenicity Cohort only), and exploratory assays 
using PBMCs, plasma, and RNA samples for correlates of risk/protection analyses in all 
subjects. 
 
ANALYSIS OF PREVENTION OF INFECTION 
The primary Mtb infection endpoint will be QFT-GIT conversion from a negative to positive 
test, using the manufacturer’s recommended threshold of 0.35 IU/mL, at any time-point after 
Day 84 and through end of follow-up for the primary endpoint. The frequency of QFT-GIT 
negativity/positivity will be summarized for each treatment group at screening, on Day 84, on 
Month 6, and for all subsequent assessments. The log-rank statistic will be used to test the null 
hypothesis of no difference in the rates of Mtb infection over the follow-up period between each 
of the AERAS-404 and BCG groups compared to the placebo group. The log-rank test statistic 
compares estimates of the hazard functions of the vaccine groups at each observed event time. It 
is constructed by computing the observed and expected number of events in the groups at each 
observed event time and then summing these to obtain an overall summary across all time points 
where there is an event. As subjects convert their QFT-GIT tests over time (experience a primary 
endpoint) or are lost to follow up the number of subjects at risk over time will change. As this is 
a proof of concept study, these two log-rank tests (AERAS-404 compared to placebo and BCG 
revaccination compared to placebo) will be performed with a 1-sided Type 1 error of 0.1. There 
will be no adjustment for multiplicity to control the Type 1 error rate over the two tests. The 
rationale for not performing this adjustment is related to the fact that the tests pertain to efficacy 
assessments of two unrelated vaccines. If these two evaluations were performed in two separate 
trials then no adjustment for multiplicity would be made. Thus, one should not be compelled to 
adjust for this multiplicity simply because the two evaluations are performed administratively in 
a single trial.  
 
The secondary Mtb infection endpoint will be the combination of QFT-GIT conversion from a 
negative to positive test, using the manufacturer’s recommended threshold of 0.35 IU/mL, at any 
time-point after Day 84 and through end of follow-up for the primary endpoint, AND persisting 
without QFT-GIT reversion from a positive to a negative test through 6 months after QFT-GIT 
conversion. The methods used for the analysis of this endpoint will be the same as that for the 
primary Mtb infection endpoint, but the number of endpoints will be smaller as subjects who 
experience primary QFT-GIT conversion but then have a reversion to negative at one or both of 
the repeat tests will not be counted. 
 
ANALYSIS OF IMMUNOLOGY 
The primary variables of interest for preliminary assessment of immune response to vaccine will 
be the percentage of CD4+ and CD8+ T cells that express IFN-γ, TNF, IL-2, IL-17, IL-22, 
CD107a, and/or CD154 alone or in combination in response to stimulation with peptide pools 
representing the entire amino acid sequence of the mycobacterial antigens Ag85B and TB10.4, 
and BCG antigens.  Due to high backgrounds associated with the CD107a response, CD107a 


 
 
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25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
expression in the absence of any other functional response will be ignored.  Response will be 
measured by flow cytometry in the intracellular cytokine staining (ICS) assay. 
 
ANALYSIS OF SAFETY 
The safety profile of BCG revaccination, AERAS-404 and saline placebo will be described by 
treatment group.  The primary variable for evaluation of the safety profile will be the number and 
percentage of unsolicited and solicited adverse events recorded at all available post-vaccination 
time points.   


 
 
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AERAS-404 
 
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25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
1 
INTRODUCTION 
There were an estimated 8.7 million new cases of tuberculosis (TB) worldwide in 2011, (13% 
co-infected with HIV) and 1.4 million people died from TB, including almost one million deaths 
among HIV-negative individuals (WHO, 2012). Although new cases of TB have been falling 
worldwide for several years and the TB mortality rate has decreased 41% since 1990, Africa is 
not on track to reach TB mortality targets by 2015. South Africa ranks 3rd among the top twenty 
high TB burden countries worldwide, with 500,000 new TB cases each year, and the highest for 
TB incidence per population (993 per 100,000). Improved control of TB among young adults 
would have significant impact on the South African TB epidemic (Dye and Williams, 2008; 
Abu-Raddad et al, 2009). The spread of multi-drug and extensively drug resistant TB also 
highlights the need for an effective vaccine against this ancient disease.  
 
Clinical development of TB vaccines is hampered by the lack of biologic correlates of protection 
or validated preclinical models, which provide evidence of likely efficacy for advancement into 
large scale trials. As an example, despite protection in some animal models and immunogenicity 
in target populations, the novel vaccine MVA85A did not offer additional protection against TB 
disease or Mtb infection in infants beyond that offered by BCG vaccination at birth (Tameris et 
al, 2013). Several additional candidate vaccines are advancing in clinical development, but the 
resources to test all of these vaccines in costly and lengthy clinical trials are likely to fall short. 
Infection with Mtb is a much more frequent event than TB disease, so the size and duration of a 
clinical trial to test for prevention of infection with Mtb is much less than that of a study testing 
for prevention against TB disease. While a TB vaccine would not need to prevent infection with 
Mtb to prevent TB disease, prevention of infection with Mtb would be an important marker of 
biologic impact, and likely efficacy, since lasting prevention of Mtb infection would interrupt the 
cycle of disease and transmission.  
 
A vaccine targeting adolescents or young adults would have maximal impact on TB control (Dye 
and Williams, 2008; Abu-Raddad et al, 2009; Dye, 2000), since TB incidence after adolescence 
is high (see Figure 1) and it is adolescents and adults who are responsible for the vast majority of 
transmission (Mahomed et al, 2011a). Thus adolescents and young adults are an important target 
population for TB vaccines. Boosting of BCG-induced immunity with novel TB vaccines, or 
with BCG revaccination, might prevent progression to active pulmonary TB disease in young 
adulthood and the cycle of transmission.  
 
Bacille Calmette Guerin (BCG) Vaccine SSI is 
registered in South Africa for prevention of TB 
in unvaccinated infants, children, and adults. 
Protection against TB is highly variable, and 
likely to be related to regional differences in 
environmental mycobacterial exposure. (Fine, 
1995). Some evidence supports the hypothesis 
that BCG vaccination offers protection against 
Mtb infection in young children (Soysal et al, 
2005; Eisenhut et al 2009; Basu et al, 2012). 


 
 
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There is also some evidence that BCG revaccination of adolescents offers modest protection in 
temperate geographical areas (Barret et al, 2011; Pereira et al 2012; Rodrigues et al, 2005). It is 
also possible that BCG revaccination of adolescents might protect against primary Mtb infection, 
but this hypothesis has yet to be tested and is one of the objectives of this study. 
 
AERAS-404 is a novel TB vaccine originally developed by SSI (Copenhagen, Denmark) and 
currently being jointly developed by SSI, Sanofi Pasteur and Aeras.  AERAS-404 is intended to 
be deployed as a booster vaccine following BCG. Initial safety and immunogenicity have been 
demonstrated in adults in several clinical trials as described below and in the Investigator’s 
Brochure (IB). While a closely related vaccine (see Section 1.5) is currently being evaluated in 
adolescents, this will be the first use of AERAS-404 in adolescents. A dose-escalating safety and 
immunogenicity trial in South African infants using AERAS-404 (IMPAACT P1113/C-015-404) 
has been approved by the Medicines Control Council and is expected to begin enrollment in 
2013. Demonstration of a biologic impact against infection with Mtb in adolescents would 
provide additional rationale for a Phase 2b study of protection against TB disease of AERAS-404 
in infants or an adolescent/young adult population.    
 
1.1 
Background  
The high incidence of Mtb infection and active TB disease among adolescents in high TB burden 
countries suggests that trials assessing vaccine-induced protection are critical in this age group. 
Approximately 50% of high school students in the Worcester region, near Cape Town, South 
Africa, test QuantiFERON-TB Gold in-tube (QFT-GIT) positive (Mahomed et al, 2011a). The 
incidence of active TB disease in this same adolescent study population is similarly high (450 
per 100,000 person years) (Mahomed et al, 2013). Further, the increase in active TB disease 
incidence that begins in adolescence occurs in parallel with a very high incidence of QFT-GIT 
conversion, with QFT-GIT conversion rates (the denominator being QFT-GIT negative 
individuals) increasing by 14% to 17% per year between 13-17 years of age (unpublished SATVI 
data).  
 
Adolescents who are QFT-GIT positive have an almost three-fold higher incidence of active TB 
disease in subsequent years, compared to QFT-GIT negative adolescents (640 vs 220 per 
100,000 person years) (Mahomed et al, 2011b). However, the highest risk of TB disease occurs 
in adolescents with recent QFT-GIT conversion. Adolescents who were known to have 
converted from QFT-GIT negative to QFT-GIT positive over a preceding two-year period had 
eight-fold higher TB disease incidence in subsequent years, compared to adolescents who had 
remained QFT-GIT negative (1,460 vs 170 per 100,000 person years) (Machingaidze et al, 
2012). Therefore, QFT-GIT conversion has clear biological importance as a risk factor for active 
TB disease in this age group. It follows that demonstration of prevention of QFT-GIT conversion 
would be a critical step towards control of TB disease incidence among young adults. This 
clinical trial will test whether vaccination with AERAS-404 or BCG revaccination prevents 
QFT-GIT conversion among healthy, HIV-uninfected, QFT-GIT negative, previously BCG 
vaccinated adolescents. 
 


 
 
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The QFT-GIT assay has limitations as a test for Mtb infection, due to intra-subject and dynamic 
variability around the diagnostic threshold recommended by the manufacturer and by the US 
Centers For Disease Control And Prevention (CDC) (0.35 IU/mL) (Pai, 2012; Mazurek et al, 
2010). In a study of short-term serial QFT-GIT testing in South Africa, QFT-GIT assays were 
associated with a 4% false positive conversion rate, supporting the recommendation that an 
“uncertainty zone” of 0.2 – 0.7 IU/mL (i.e., increasing the positive threshold value) be applied to 
interpretation of QFT-GIT results (van Zyl-Smit et al, 2009). QFT-GIT assay variability appears 
maximal in low burden settings with low rates of TB infection. Repeat testing of individual QFT-
GIT samples in a low burden setting showed that the normal expected range of within-subject 
variability is ±0.24 IU/mL (coefficient of variation 27%) for individuals with a QFT-GIT 
response within this “uncertainty zone” (Metcalfe et al, 2013). Serial QFT-GIT testing of 
Canadian health care workers, with expected TST conversion rates of less than 1%, has also 
shown unexpectedly high QFT-GIT conversion rates (5%) unrelated to occupational TB 
exposure risk (Zwerling et al, 2013). These unexpected QFT-GIT conversions were associated 
with a high rate of spontaneous reversion (62%).  
 
 
Findings from high TB burden settings with high risk of Mtb infection, such as South Africa, 
where the rate of QFT-GIT conversion is similar to the expected rate of TST conversion, are in 
contrast to the pattern observed in serial testing of low risk study populations, where the rate of 
QFT-GIT conversion has been unexpectedly high compared to TST (Pai, 2012; Zwerling et al, 
2013). For example, in South African adolescents, the QFT-GIT assay performs equally as well 
as the TST for prediction of subsequent active TB disease (Mahomed et al, 2011b). In a high risk 
South African community, the QFT-GIT conversion rate among QFT-GIT baseline negative 
adolescents was less than the rate of TST conversion (5mm threshold) over the same period 
(17% vs. 24%) (unpublished SATVI data). These preliminary data indicate that QFT-GIT 
conversion is an appropriate endpoint to define Mtb infection, and vaccine-induced prevention of 
Mtb infection, for clinical trials among South African adolescents. 
 
Optimum use of QFT-GIT conversion as an infection end-point requires further exploration of 
the effect of alternative definitions of positive and negative test results on rates of QFT-GIT 
conversion, reversion, and protection in multiple populations and in high and low burden 
countries. These alternative conversion thresholds will be evaluated as exploratory objectives in 
this clinical trial. 
 
1.2 
Description of AERAS-404 Vaccine  
AERAS-404 has two components: the H4 antigen and the IC31 adjuvant.  The term HyVac4 
refers to the H4 antigen and IC31 adjuvant in non-clinical development by SSI. The term 
AERAS-404 refers to the H4 antigen and IC31 adjuvant in clinical development sponsored by 
Aeras (Rockville, Maryland USA).  The current formulation of AERAS-404 is a field-
reconstituted vaccine with H4 antigen and IC31 adjuvant supplied in different vials. The 
components are dissolved in a sterile aqueous buffer containing tris-
hydroxymethylaminomethane (Tris) and sodium chloride (NaCl). 
 


 
 
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H4 Antigen 
The H4 antigen is a fusion protein created from two Mtb antigens: antigen 85B (Ag85B) and 
TB10.4. Ag85B is also referred to as α-antigen and is a 30-kDa mycolyl transferase protein 
(Horwitz et al, 2000; Belisle et al, 1997).  TB10.4 is one of three members of the very similar 
ESAT-6 group of proteins found in Mtb culture supernatants. TB10.4 induces broad immune 
responses in T cells isolated from TB subjects compared to BCG-vaccinated donors and 
unvaccinated donors (Skjot et al, 2000; Skjot et al, 2002).  
 
IC31 Adjuvant   
The IC31 adjuvant is a proprietary adjuvant of Intercell AG (Intercell, Vienna, Austria). IC31 is 
a combination of a leucine-rich peptide, named KLK, and a synthetic oligonucleotide, named 
ODN1a.  KLK enhances the uptake of antigens into the antigen-presenting cell and increases the 
immune response to peptide antigens. ODN1a is a synthetic bacterial deoxyribonucleic acid 
(DNA) analogue that resembles a CpG pattern that will direct the adaptive immune response 
toward a T helper cell type-1 (Th-1) pattern with production of interferon-γ (IFN-γ) and 
interleukin-12 (IL-12). The amount of adjuvant given may affect the immune response. The 
optimal molar ratio of KLK to ODN1a in mice is 25:1 (Lingnau et al, 2002).  This same molar 
ratio is used in all formulations of AERAS-404 tested in clinical studies.  For simplicity and 
clarity hereafter, amounts and concentrations of IC31 adjuvant will be expressed as molar 
equivalents of KLK. 
 
1.3 
Nonclinical Experience with AERAS-404 (HyVac4) Vaccine 
Multiple toxicology studies have been conducted in mice and rabbits, in both BCG-primed and 
non-primed animals. Administration of HyVac4 was not associated with overt signs of toxicity 
and did not demonstrate mortality, adverse clinical signs, effects on body weight or food 
consumption, body temperature, ophthalmology, or clinical pathology.  
 
The H4 antigen used in AERAS-404 has been evaluated in mice at 0.5, 5.0 and 15 mcg, in 
guinea pigs at 20 mcg, and in rabbits at 150 mcg in combination with IC31 adjuvant.  HyVac4 is 
immunogenic in mice and induced a significant additive protective efficacy against subsequent 
aerosol challenge with Mtb compared to BCG alone (Billeskov et al, 2012; Skeiky et al, 2010; 
Dietrich et al, 2005).  Administration of 20 mcg of H4 antigen after BCG prime has been shown 
to protect guinea pigs from aerosol challenge with Mtb compared to BCG alone (Skeiky et al, 
2010).  Ag85B has been previously demonstrated in the guinea pig model to induce substantial 
protective immunity against aerosol challenge with the highly virulent Erdman strain of Mtb 
(Horwitz et al, 2000).   
 
Additional details regarding the nonclinical experience and results of toxicology studies with 
HyVac4 can be found in the IB.   
 
1.4 
Clinical Experience with AERAS-404 
AERAS-404 has been studied in 198 adults who were enrolled in 4 Phase I studies as shown in 
Table 1-1; an additional 36 subjects received placebo. Subjects received 2 or 3 doses of the 
vaccine or placebo at the doses indicated. Three studies were conducted in European adults (a 


 
 
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TB non-endemic population) and one was conducted in a TB endemic population (South Africa). 
Subjects were previously BCG-vaccinated, either in childhood or as part of a prime-boost 
regimen. All were QuantiFERON (QFN) negative (non-latently infected) prior to enrollment. 
 
Table 1-1 
Clinical Studies of AERAS-404 Performed to Date 
Study 
Number 
Population/Location 
Treatment regimen 
H4(μg)/IC31(nmol) 
Number of doses/ 
Dosing schedule 
Number receiving 
vaccine 
C-005-404 
18-45y / Sweden 
BCG pos, QFN neg, 
HIV neg 
50,150 / 0,100,500 
1 or 2 / SD 0, 56 
56 
(24 received H4 
alone) 
C-006-404 
18-45y / Finland 
BCG pos, QFN neg, 
HIV neg 
5,15,50,150 / 100,500 
2 / SD 0, 56 
50 
C-011-404 
18-45y / South Africa 
BCG pos, QFN neg, 
HIV neg 
5,15,50,150 / 500 
2 / SD 0, 56 
32 
C-013-404 
18-45y / Switzerland 
BCG neg, QFN neg, 
HIV neg 
(BCG prime at SD -42)  
50 / 500 
3 / SD 0, 56, 231 
or 
2 / SD 56, 231 
60 
 
No vaccine-related serious adverse events have occurred and the vaccine has generally been well 
tolerated at all doses and regimens evaluated. Injection site reactions (pain, swelling, and 
erythema) have been mild or moderate. Reactions have occurred at the site of tuberculin skin 
tests (TST) and BCG vaccination sites as described below. Systemic adverse reactions include 
mild to moderate fatigue, myalgia, headache, arthralgia, and mild to severe pyrexia. 
Asymptomatic transient isolated proteinuria was seen in subjects receiving the vaccine and 
placebo in all four trials and was more frequent in South African adults, including at baseline 
(prior to vaccination). Proteinuria did not recur with revaccination and this finding is not 
considered to be clinically significant. One subject had a reactivation of Graves’ disease; and one 
subject was diagnosed with celiac disease at the end of the study; neither event was considered 
related to study vaccine.  
 
In study C-005-404 a TST result less than 10 mm was an eligibility requirement. AERAS-404 
was shown to induce post-vaccination hypersensitivity reactions (HSR) at the sites of recently 
administered TSTs, with 14/21 (66.7%) subjects who received AERAS-404 after TST 
experiencing a post-vaccination TST site HSR, characterized by warmth, erythema and 
induration, with onset from several hours to two days after Study Day 0 vaccination, and 
duration from one to 28 days, typically resolving without treatment. Two of the 14 subjects also 
experienced pruritus at the TST site. For this reason, the TST was removed as a study procedure 
from C-005-404 and subsequent protocols.   
 
Local BCG injection site reactions are well documented adverse events in subjects who receive 
BCG. Induration appears at the site of the BCG injection which is followed by a local lesion 
(redness and swelling) that may ulcerate some weeks later. In study C-013-404 all study 
participants received BCG at Study Day -42, before receiving AERAS-404 or placebo at Study 
Days 0, 56, and 231. Redness was the most common local BCG injection site reaction with all 


 
 
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subjects (100%) experiencing redness after BCG vaccination. Redness persisted in the majority 
of the subjects throughout the study, with at least 60% of subjects in each group having redness 
at the final study assessment. Swelling was the second most common local BCG injection site 
reaction. None of the redness or swelling at the BCG injection site met protocol-specified 
toxicity grading scale criteria for a severe (Grade 3 or higher) adverse event. BCG injection site 
ulceration occurred after BCG vaccination in both placebo and AERAS-404 recipients. There 
was no dose-related increase in frequency of BCG site ulceration after receiving AERAS-404. 
Overall, there was similar distribution of BCG local reactions in subjects who received AERAS-
404 and placebo, with no dose response of BCG site reactions noted in subjects who received 2 
doses or 3 doses of AERAS-404 and no evidence of acute worsening of BCG site reactions after 
AERAS-404 administration. 
 
Table 1-2 presents the number and percentage of subjects in all four Phase 1 studies of AERAS-
404 experiencing adverse events related to AERAS-404 for events occurring in ≥5% of subjects.   
 
Table 1-2 
Adverse Events in the AERAS-404 Treatment Groups in Adults with 
Event Rate of ≥5% and Number of Subjects for Each Event 
Related Adverse Eventa 
MedDRA Preferred Termb 
AERAS-404 
(N=198)c 
nd (%) 
Injection site pain 
109 ( 55.1) 
Fatigue 
95 ( 48.0) 
Headache 
82 ( 41.4) 
Myalgia 
76 ( 38.4) 
Injection site swelling 
39 ( 19.7) 
Injection site erythema 
37 ( 18.7) 
Arthralgia 
28 ( 14.1) 
Pyrexia 
25 ( 12.6) 
Neutrophil count decreased 
18 (  9.1) 
Application site hypersensitivitye 
14 (  7.1) 
Chills 
13 (  6.6) 
Diarrhoea 
12 (  6.1) 
Protein urine 
12 (  6.1) 
Red blood cells urine 
12 (  6.1) 
Haemoglobin decreased 
11 (  5.6) 
Nausea 
10 (  5.1) 
a. 
Adverse event data through the end of the study for C-005-404, C-006-404, C-011-404, and C-013-404; 
includes adverse events that occurred on or after the first dose of AERAS-404 
b. For the purpose of summarizing cumulative data across multiple studies, adverse events were coded using the 
same version of MedDRA, which may differ from the version of MedDRA that was used to code adverse 
events in an individual study. 
c. 
N is number of subjects who received at least 1 dose of AERAS-404 (HyVac4) 
d. n is number of subjects with at least 1 related adverse event for the preferred term in question 
e. 
Application site hypersensitivity refers to reactions at the tuberculin skin test application site 
 
A more detailed description of post-vaccination adverse events on all AERAS-404 clinical trials 
in adults can be found in the IB. As noted, while an infant study is expected to begin enrolling in 


 
 
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2013, and while a related vaccine is currently being evaluated in adolescents, this will be the first 
use of AERAS-404 in adolescents. 
 
Immune responses in the previous adult trials have been used to determine the dose regimen to 
be used in this study. Among participants in study C-006-404, increases in ELISpot responses 
were seen among all groups receiving AERAS-404, and peaked at Day 84. Responses were 
strongest among participants receiving IC31 at 500 nmol, with comparable responses between 
groups receiving 5, 15, and 50 mcg H4. ICS responses showed Ag85B-specific cluster of 
differentiation 4 (CD4+) T-cells among all treatment groups, peaking at 2 or 4 weeks after the 
second dose of AERAS-404. The highest magnitude of responses was noted among the 5, 15, 
and 50 mcg H4 dose groups. CD4+ responses were predominantly of bifunctional or 
polyfunctional profile. By comparison, TB10.4-specific CD4+ responses were lower in 
frequency and of lower magnitude compared to Ag85B-induced CD4 responses. 
 
Samples from participants in C-011-404 were evaluated following stimulation with Ag85B.  
Responses were noted in each dose group by 7-color Aeras ICS, with the highest responses being 
noted in the 15/500 H4:IC31 group predominantly against the Ag85B peptide pool. No new T-
cell responses were noted among participants receiving the highest level of antigen, 150 mcg H4.  
Mtb-specific CD4 T-cell responses did not significantly change in the 5/500 and 50/500 H4:IC31 
groups following the second dose of study vaccine, whereas a significant increase of Mtb-
specific CD4 T-cell responses was observed in the 15/500 H4:IC31 group (mean 0.237%) at 
D84/182 as compared to both baseline (P<0.0001) and D14/28 (P=0.0364). Responses were 
more highly polyfunctional in the 5/500 and 15/500 H4:IC31 groups.  
 
In Study C-013-404, the dosage studied, 50/500 H4:IC31, was selected before 15/500 H4:IC31 
was found to induce a more favorable immune profile in adults.  Two or three doses of AERAS-
404 boosted recent BCG priming and induced Ag85B antigen-specific CD4 T cells above that 
seen with placebo control. These responses were primarily polyfunctional (IFN-γ, IL-2 and TNF-
α) and bi-functional (IL-2 and TNF-α). In subjects receiving 3 doses of AERAS -404, the 
magnitude of Ag85B-specific CD4 T cell responses on Study Day 259 (28 days after the third 
dose) was comparable to that seen on Study Day 84 (28 days after the second dose).  The three 
dose regimen was seen to only slightly outperform the two dose regimen as measured by the 
magnitude of Ag85B-specific CD4 T-cell responses.  
 
In summary, CD4 responses have consistently been induced by vaccination with AERAS-404. 
Responses were higher after two vaccinations of the 15/500 dose compared to the higher and 
lower doses, and did not significantly increase with a third vaccination. Based on these data, a 
dose of 15/500 H4:IC31 administered on a 0, 56 Day schedule has been selected as most 
appropriate for this trial in adolescents.  
 
1.5 
Clinical Experience with Vaccines Related to AERAS-404  
Two fusion protein vaccines using similar antigens and the same adjuvant used in AERAS-404 
are also in clinical development. 
  


 
 
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AERAS-456 (H56:IC31) 
AERAS-456 is a fusion protein of the Ag85B, ESAT-6, RV2660 antigens formulated with IC31 
adjuvant, and is being jointly developed by Aeras and SSI. One previous Phase 1 trial (C-032-
456) has been conducted in South African adults. Twenty-five BCG-vaccinated adults were 
enrolled in 3 groups stratified by baseline QuantiFERON TB Gold status (latently infected) and 
H56/IC31 dosage as follows: 8 healthy subjects not infected with Mtb (dose 50/500 H56:IC31), 8 
Mtb infected subjects (dose 15/500 H56:IC31), and 9 Mtb infected subjects (dose 50/500 
H56:IC31). Three doses were administrated in all three groups at intervals of 56 days. Subjects 
were followed up for 210 days. 
  
No vaccine related SAEs and no severe AEs were reported. The most common systemic adverse 
events were headaches, proteinuria and upper respiratory tract infections with most judged not to 
be vaccine related. All local AEs were mild to moderate and transient without medical 
interventions. AERAS-456 was immunogenic in BCG-vaccinated adults with and without latent 
infection. The vaccine induced a dominant CD4+ T cell responses dominated by polyfunctional 
CD4+ T cells. A greater boosting effect was observed in LTBI (+) subjects than in LTBI (-) 
subjects. 
   
Hybrid-1:IC 
The Hybrid-1:IC vaccine is being developed by SSI, and consists of the Hybrid 1 fusion protein 
of the Mtb antigens ESAT-6 and Ag85B in combination with IC31.  Three Phase I clinical trials 
in healthy adults have been completed. These studies enrolled a total of 95 healthy, HIV (-) 
adults with and without latent TB infection, 73 of whom received up to 2 vaccinations with 50 
mcg antigen and 100 or 500 nmol of IC31 (van Dissel et al, 2010; van Dissel et al, 2011). A 
Phase II trial evaluated two vaccinations of 40 HIV infected adults with CD4 counts >350, and 
was completed in 2013.  Another Phase II trial in South African adolescents with and without 
latent TB infection is currently enrolling; a total of 240 subjects will receive 1 or 2 doses of 15 or 
50 mcg antigen and 500 nmol of IC31.  
  
Two possible vaccine related SAEs were observed (elevated CPK and AST/ALT values, not 
associated with clinical abnormalities) but the Hybrid-1:IC vaccine has generally been well 
tolerated, with most adverse events mild or moderate. No safety concerns have been identified, 
including in populations with HIV infection and latent TB infection, and in adolescents. 
 
1.6 
Description of BCG 
BCG Vaccine SSI is manufactured by Statens Serum Institut (SSI), Copenhagen, Denmark and 
imported and distributed in South Africa for the national immunization program by Biovac, 
Johannesburg, South Africa. BCG Vaccine SSI is registered in South Africa for prevention of TB 
in children and adults. BCG, an attenuated, live culture of the Bacillus Calmette-Guérin, was 
originally attenuated between 1906 and 1919 by serial passage of an M. bovis strain. The Statens 
Serum Institut in Copenhagen, Denmark derives this vaccine from the Danish BCG strain 1331. 
SSI BCG is supplied by the manufacturer in amber 10-dose vials containing 0.75 mg lyophilized 
SSI BCG. The vaccine will be reconstituted with Sauton SSI diluent supplied by the 
manufacturer, according to the manufacturer’s instructions. After reconstitution, 1 adult dose 
(0.1 mL) contains 2 to 8 x 105 CFU. Sauton SSI Diluent is composed of magnesium sulphate, 


 
 
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dipotassium phosphate, citric acid, monohydrate L-asparagine monohydrate, ferric ammonium 
citrate and glycerol at 85%, which is reconstituted with sterile water for injection. 
 
There is considerable experience with BCG vaccine, which is licensed in South Africa for 
prevention of TB in children and adults. Local and systemic reactogenicity in various 
populations is well described (FitzGerald 2000; Hoft et al 1999; Lotte et al 1984; Nicol et al 
2002). Injection site and regional complications, such as extensive local ulceration, local 
subcutaneous abscesses, and suppurative lymphadenitis, occur in less than 1 per 5 million 
vaccinations. Induration and redness of the skin at the site of intradermal BCG vaccination 
typically develops within several days and represents a normal response to BCG vaccination. 
The induration gradually resolves over several days and is followed by a small local superficial 
ulcer. The ulcer opens and drains for 4 weeks on average, and spontaneously heals within 2-3 
months, usually leaving a small scar. A brief period of minor, asymptomatic enlargement of the 
regional cervical and axillary lymph nodes (< 1 cm) is common. Systemic adverse events due to 
BCG occur very infrequently. Fever, headache and non-injection site cutaneous manifestations 
occur in less than 1% of vaccinees. Severe systemic adverse events, such as osteitis and 
disseminated BCG disease, are rare (about 1 per 5 million vaccinations) and usually occur in 
immune-compromised infants. Severe local or systemic complications of BCG would not be 
expected in the healthy, HIV-uninfected, Mtb uninfected adolescents to be recruited in this trial. 
 
Adolescents in this trial will have received prior BCG vaccination at birth. BCG revaccination 
appears to be safe and well tolerated in adolescents and young adults (Pereira et al 2012; 
Rodrigues et al, 2005; Bottiger et al 1983). In a large trial of BCG revaccination in more than 
7,000 Brazilian adolescents, only 25 of the participants had adverse reactions and no deaths or 
cases of disseminated BCG disease were reported (Pereira et al 2012; Rodrigues et al, 2005). 
Other published trials of BCG revaccination have concluded that BCG revaccination is not 
associated with significantly more AEs than primary BCG vaccination. In a trial of BCG 
revaccination in 2,997 fourteen to fifteen year old Swedish adolescents, open vaccination lesions 
were reported in 4% (mean diameter of open lesions 4mm) of adolescents receiving the Danish 
BCG vaccine that will be used in this trial (Bottiger et al 1983). South African Tuberculosis 
Vaccine Initiative (SATVI) data from an ongoing trial of BCG revaccination and isoniazid 
preventive therapy (South African National Clinical Trials Register DOH-27-0212-3995) 
indicate that BCG revaccination is safe and well tolerated in young adults. Eighty-two subjects in 
this ongoing trial were enrolled with a TST ≥15mm. Data from these Mtb-infected young adults 
show that injection site reactions are mostly of mild intensity, with ulceration of the injection site 
resolving within three months. No BCG-related SAEs or major safety concerns have arisen.  
 
1.7 
Rationale for Study 
This clinical trial will evaluate safety, immunogenicity, and prevention of Mtb infection (QFT-
GIT conversion), of AERAS-404 and BCG revaccination in healthy, HIV-uninfected, QFT-GIT 
negative, previously BCG vaccinated adolescents. QFT-GIT negative adolescents in the area 
where this study will be conducted have previously been shown to have very high rates of TST 
and QFT-GIT conversion, associated with increased risk of active TB disease in subsequent 
years (Mahomed et al, 2011b; Machingaidze et al, 2012). A TB vaccination strategy 
incorporating BCG revaccination or AERAS-404 vaccination in adolescents or young adults, if 


 
 
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found to prevent Mtb infection, would likely have a major impact on TB disease, TB 
transmission, and control of the epidemic (Dye and Williams, 2008; Abu-Raddad et al, 2009; 
Dye, 2000). If revaccination with BCG or vaccination with AERAS-404 is shown to prevent 
infection with Mtb in this proof of concept study in adolescents, additional larger scale studies 
examining the impact on TB disease in more diverse populations would be warranted. 
 
2 
STUDY OBJECTIVES AND DESIGN  
2.1 
Objectives 
Primary Objectives:  
• To evaluate the safety profile in HIV-uninfected, remotely BCG vaccinated adolescents of 
o AERAS-404 
o BCG revaccination 
• To evaluate prevention of Mtb infection, as measured by rates of conversion using a QFT-
GIT assay, by 
o AERAS-404 compared to placebo 
o BCG revaccination compared to placebo 
 
Secondary Objectives: 
• To evaluate prevention of Mtb infection, as measured by rates of sustained conversion using 
a QFT-GIT assay, by  
o AERAS-404 compared to placebo 
o BCG revaccination compared to placebo 
• To investigate the immunogenicity in HIV-uninfected, remotely BCG vaccinated adolescents 
of 
o AERAS-404 
o BCG revaccination 
 
As this is a novel study design, generation of data and hypotheses to be tested in future vaccine 
trials is an important overarching goal. Thus the exploratory objectives below will also be 
investigated: 
• To evaluate prevention of Mtb infection, as measured by rates of reversion to a negative from 
a positive QFT-GIT assay, by 
o AERAS-404 compared to placebo 
o BCG revaccination compared to placebo  
• To explore the effect of alternative QFT-GIT test threshold values on rates of QFT-GIT 
conversion, QFT-GIT reversion, and prevention of Mtb infection 
• To identify immune correlates of risk for Mtb infection  
• To identify immune correlates of vaccine-induced protection against Mtb infection induced 
by AERAS-404 or BCG revaccination 
• To evaluate alternative IGRAs or immune markers for diagnosis of Mtb infection or as 
markers of exposure to non-tuberculous mycobacteria (NTM) 
• To explore trends in TB disease incidence after AERAS-404 vaccination or BCG 
revaccination 


 
 
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• To explore trends in QFT-GIT prolonged/sustained conversions and late reversions (i.e., 
more than 6 months post initial conversion) in early QFT-GIT converters (i.e., among those 
who converted at Month 6 or Month 12 of follow-up) 
 
2.2 
Design and Endpoints 
This Phase II, randomized, 3-arm, placebo controlled, partially blinded, clinical trial will be 
conducted in 990 healthy, HIV-uninfected, QFT-GIT negative, previously BCG vaccinated 
adolescents. The trial will be conducted at the SATVI site in the Worcester region of the Western 
Cape region of South Africa, where epidemiological studies involving thousands of adolescents 
have been conducted over the last decade to characterize rates of Mtb infection and active TB 
disease in this age group (Mahomed et al, 2011b; Mahomed et al, 2013). Additional sites with 
similar TB epidemiology may also be used. 
 
Subjects will be recruited from high schools or directly from the community. The all-TB disease 
incidence in the Worcester region is greater than 1% per year (2011 DOH data). TB disease 
incidence in 12-18 year old adolescents attending high school is 450 per 100,000; approximately 
40% of adolescents are Mtb infected at entry to high school; and HIV prevalence among 
adolescents is less than 1% (Mahomed et al, 2011b; Mahomed et al, 2013). 
 
Subjects will be randomized in a 1:1:1 ratio to receive either two doses of AERAS-404 (15 mcg 
H4/500 nmol IC31), or two doses of placebo at equivalent volume, administered by 
intramuscular injection on Day 0 and Day 56; or one dose of BCG Vaccine SSI (2-8 x 105 CFU) 
administered by intradermal injection on Day 0. Allocation to the AERAS-404 and saline 
placebo trial arms will be double-blinded. Since BCG causes a recognizable local injection site 
reaction, the BCG revaccination trial arm will be unblinded. Laboratory staff performing QFT-
GIT, immunogenicity, and correlates assays will be blinded to all three interventions.  
 
Subjects will be enrolled in two sequential cohorts and within each cohort subjects will be 
randomized in a 1:1:1 ratio to receive either AERAS-404, saline placebo, or BCG Vaccine SSI. 
The first 90 subjects (30 from each arm) will form the Safety & Immunogenicity Cohort and will 
be subject to more intensive collection of safety data, with data reviewed by the Data Monitoring 
Committee (DMC), principal investigator and local medical monitor. Selected immunogenicity 
assays, including whole blood intracellular cytokine staining (ICS), will also be performed in this 
cohort. The remaining 900 subjects will be enrolled into the Correlates Cohort.  All 990 subjects 
in the study will be evaluated for safety and biomarker outcomes, and for prevention of Mtb 
infection. 
 
Allocation of subjects by treatment group and study cohort is shown in Table 2-1.  
 
  


 
 
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Table 2-1 
Distribution of Subjects by Treatment Group and Study Cohort 
 
 
Study Cohort 
Planned Number of Subjectsa 
Treatment Group 
 
Total 
Placebo 
BCG 
AERAS-404 
Safety & Immunogenicity 
30 
30 
30 
90 
Correlates 
300 
300 
300 
900 
Total 
330 
330 
330 
990 
a. 
Subject numbers are approximate due to block randomization. 
 
Safety outcomes will include solicited adverse events (AEs) reported through 7 days after each 
vaccination and unsolicited adverse events reported through 28 days after each vaccination, 
injection site assessments performed through Day 84, and adverse events of special interest 
(AESIs), serious adverse events (SAEs), and suspected unexpected serious adverse reactions 
(SUSARs) reported until end of study. The Data Monitoring Committee (DMC) will review 
unblinded safety data from the Safety & Immunogenicity Cohort twice: data through Day 7 and 
data through Day 84. The DMC may request additional information, or a pause in recruitment 
and vaccination, while safety data are being evaluated. The DMC will make a formal 
recommendation on the continued conduct of the trial after each safety review; if pausing rules 
have not been met enrollment will continue while the reviews are being conducted. Each subject 
will be followed for safety for a minimum of 6 months after the last vaccination. 
 
The primary Mtb infection endpoint will be QFT-GIT conversion from a negative to positive 
test, using the manufacturer’s recommended threshold of 0.35 IU/mL, at any time-point after 
Day 84 and through end of follow-up for the primary endpoint (see Figure 2-1) (Mazurek et al, 
2010). The 84-day ‘wash-out’ period is stipulated in order to exclude subjects who may have 
already been Mtb infected but not yet converted their QFT-GIT test at screening, thus subjects 
who convert their QFT-GIT at Day 84 will not be included in the analyses of prevention of Mtb 
infection.   
 
The duration of the trial will be endpoint-driven, using a design targeting accrual of 64 primary 
endpoints. The primary analysis will be triggered when at least 64 endpoints are accrued in the 
study (both cohorts) AND the median follow up time is at least 15 months, with a maximum 
individual follow-up time to detect initial QFT-GIT conversion of 24 months. Enrolment is 
expected to take approximately 12 months and the primary analysis is expected to occur 
approximately 21 months from the first subject enrolled. Therefore, duration of individual 
subject follow-up for the primary analysis is expected to range from 9 – 21 months (maximum 
24 months), with 6 months of additional follow up for those who convert to QFT-GIT positive 
after Day 84 as shown in Figure 2-1. If an initial reduction in infection rate in the BCG or 
AERAS-404 groups is seen in the primary analysis of QFT-GIT conversion, follow up for the 
entire study may be extended to examine the duration of prevention of Mtb infection. 
 
The trial sample size of 990 subjects is determined by the primary prevention of Mtb infection 
objective (measured by prevention of conversion to a positive QFT-GIT test). A potential 
mechanism of action of vaccination might be to allow initial Mtb infection (and QFT-GIT 
conversion), but to prevent establishment of sustained latent Mtb infection (and thus persistent 


 
 
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QFT-GIT conversion), resulting in QFT-GIT reversion to negative. Thus the study will also 
examine the persistence of QFT-GIT conversion.  
 
All subjects with primary QFT-GIT conversion will be followed for an additional 6 months post-
conversion to ascertain the sustained QFT-GIT conversion and QFT-GIT reversion endpoints. 
Subjects with an initial QFT-GIT conversion at Month 6 or 12 will be asked to return for a final 
QFT-GIT evaluation and assessment for TB signs and symptoms at least 24 months after their 
initial vaccination. The secondary Mtb infection endpoint will be the combination of QFT-GIT 
conversion from a negative to positive test, using the manufacturer’s recommended threshold of 
0.35 IU/mL, at any time-point after Day 84 and through end of follow-up for the primary 
endpoint, AND persisting without QFT-GIT reversion from a positive to a negative test through 6 
months after QFT-GIT conversion. The exploratory Mtb infection endpoints will be defined by 
alternative threshold values for QFT-GIT conversion and reversion (Pai, 2012; van Zyl-Smit et 
al, 2009; Metcalfe et al, 2013), alternative time periods for definition of persistent QFT-GIT 
conversion, and the evaluation of novel IGRAs which are currently being developed; details on 
these endpoint cut-off values will be included in the statistical analysis plan.  
 
The secondary analysis will be triggered after 6 months of additional post-conversion follow-up 
for individuals who convert, regardless of the rate of QFT-GIT reversion. Thus, total duration of 
the trial is expected to be 27-30 months (21-24 months, plus 6 months additional follow up for 
converters), with a maximum total duration of 42 months (12 months enrollment, 24 month 
maximum individual follow up, plus 6 months additional follow up for converters). The schedule 
of follow up for each individual subject is contingent on results of the QFT-GIT tests conducted 
at Month 3 (Day 84), and Months 6, 12, 18, and 24. For example, a subject who has negative 
QFT-GIT tests at screening, Month 3, and Month 6, but who has a positive QFT-GIT at Month 
12, will then have two additional visits with QFT-GIT testing at Months 15 and 18, at which 
time follow up will be complete for that subject. A schematic of follow up and QFT-GIT testing 
is shown in Figure 2-1.  
 


 
 
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Figure 2-1 
Schematic of Follow Up and QFT-GIT Testing 
 
Based on previous data from baseline QFT-GIT negative adolescents at the SATVI site, we 
expect the incidence of active TB disease in the study population to approximate 0.2% (estimate 
2 cases per year of follow-up) (Mahomed et al, 2011b). While the total number of active TB 
disease cases is expected to be very low and the study is not powered to detect a difference in 
rates of active TB disease among the vaccine groups, data on the number and type of cases of 
active TB disease (as defined in Section 3.6.1) will be captured as part of the assessment of 
safety.  
 
Immunological outcomes will include intracellular cytokine staining (ICS) on PBMCs and whole 
blood, transcriptomics on RNA from whole blood, absolute blood cell subset counting by flow 
cytometry (for deconvolution of transcriptomics), quantification of soluble immune mediators by 
multiplex ELISA (all in the Safety & Immunogenicity Cohort only), and exploratory assays 
using PBMCs, plasma, RNA, and absolute blood cell subset counting samples for correlates of 
risk/protection analyses in all subjects. 
 
3 
STUDY PROCEDURES  
3.1 
Schedule of Subject Evaluations  
A Summary Schedule of Evaluations depicting visit-specific procedures is provided in Table 3-1 
for subjects receiving BCG and Table 3-2 for subjects receiving AERAS-404 or placebo. Written 
informed consent and assent must be obtained before any screening procedures are performed. 
See Appendix A for a more detailed description of the evaluations.  
 


 
 
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Table 3-1 
Schedule of Subject Evaluations (Subjects receiving BCG, N=330) 
Study Visit Day (D) or Month (M)  
Screen 
0 
D7 
D28 
D70 
D84c 
M6 (D168)d,e 
M12 (D336) 
M18 (D504) 
M24 (D672) 
Eligibility criteria verification 
X 
X 
 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
Physical examination 
X 
 
 
 
 
 
 
Urine HCG (all females) 
X 
X 
 
 
 
 
 
QuantiFERON-TB Gold in-tube (mL) 
3 
 
 
 
 
3 
3 
HIV-1 (mL) with HIV counselling  
3 
 
 
 
 
 
 
Urinalysis 
X 
 
Xb 
 
 
 
 
Serum chemistry (mL)g 
5 
 
5b 
 
 
 
 
CBC, differential, platelets (mL) 
5 
 
5b 
 
 
 
 
Vital signs 
X 
X 
 
 
 
 
 
Interval history 
 
X 
X 
X 
X 
X 
X 
Focused physical examinationa 
 
X 
X 
X 
X 
X 
X 
Distribute/review diary cards 
 
Xb 
Xb 
 
 
 
 
Solicited adverse events (incl. con. meds.) 
 
X 
X 
 
 
 
 
Unsolicited adverse events (incl. con. meds.) 
 
X 
X 
X 
 
 
 
SAEs, AESIs, and SUSARs (incl. con. meds.) 
 
X 
X 
X 
X 
X 
X 
Solicited and unsolicited injection site reaction 
adverse events (incl. con. meds.)  
 
X 
X 
X 
X 
X 
 
Site of injection examination 
 
X 
X 
X 
X 
X 
 
TB symptom screen 
X 
 
 
X 
X 
X 
X 
BCG administration 
 
X 
 
 
 
 
 
Whole blood assay (mL) 
 
6b 
 
 
6b 
 
 
Absolute blood count (mL) 
 
0.5 
0.5 
 
 
 
0.5 
RNA (mL) 
 
2.5 
2.5 
 
 
 
2.5 
PBMC for ICS  (mL) 
 
8b 
 
 
8b 
 
 
Serum for multiplex ELISA (mL) 
 
3b 
3b 
 
 
 
 
PBMC, plasma, and serum for correlates of 
risk/protection (mL) 
 
16 
 
 
16 
 
40i 
Whole blood for evaluation of novel IGRA (mL)f 
7 
7 
 
 
 
7 
7 
PBMC for evaluation of NTM exposure (mL) 
 
10 
 
 
 
 
 


 
 
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Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
Study Visit Day (D) or Month (M)  
Screen 
0 
D7 
D28 
D70 
D84c 
M6 (D168)d,e 
M12 (D336) 
M18 (D504) 
M24 (D672) 
Per visit phlebotomy volume (mL)h 
23 
53 
16 
0 
32.5 
10 
53 (M6,M12)/ 
13 (M18,M24) 
Cumulative phlebotomy volume (mL)h 
23 
76 
92 
92 
124.5 
134.5 
266.5  
a. 
If indicated by interval history 
b. Subjects in the Safety & Immunogenicity Cohort only 
c. 
Subjects who convert from QFT-GIT(-) to (+) at Day 84 will have a final visit at 168 days ±14 days after 
vaccination for assessment of SAEs, AESIs, SUSARs, and TB symptom screen 
d. Month 6, 12, 18, and 24 study visits will be completed by subjects who did not convert from QFT-GIT (-) to (+) 
at the previous visit; the Month 24 visit will be the final visit for subjects who remain QFT-GIT(-) throughout 
the study.  
e. 
Subjects who convert from QFT-GIT (-) to (+) at Month 6-24 visits will have two additional visits with repeat 
QFT-GIT testing, assessment of SAEs, AESIs, SUSARs, and TB symptom screen 84 days ±14 days and 168 
days ±14 days after conversion. Up to 10 mL of blood will be collected at each of those visits, including blood 
for absolute blood count and RNA. Subjects who initially convert to QFT-GIT(+) at Month 6 or 12 will be asked 
to have a final QFT-GIT test, blood collection for absolute blood count and RNA, and TB symptom screen done 
at least 24 months after their initial vaccination.  
f. 
May not be collected at all time points shown. 
g. Serum chemistry includes AST, ALT, alkaline phosphatase, total bilirubin, creatinine, and BUN. 
h. Blood volumes are approximate. 
i. 
Study Months 6 and 12 only. 
 
Table 3-2 
Schedule of Subject Evaluations (Subjects receiving AERAS-404 
[N=330] or placebo [N=330]) 
Study Visit Day  
(D) or Month (M)  
Screen 
0 
D3  
D7 
D28 
D56 
D63 
D70 
D84c  M6  (D168)d,e 
M12 (D336) 
M18 (D504) 
M24 (D672) 
Eligibility criteria verification 
X 
X 
 
 
 
X 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
 
 
 
Physical examination 
X 
 
 
 
 
 
 
 
 
 
Urine HCG (all females) 
X 
X 
 
 
 
X 
 
 
 
 
QuantiFERON-TB Gold in-
tube (mL) 
3 
 
 
 
 
 
 
 
3 
3 
HIV-1 (mL) with HIV 
counselling  
3 
 
 
 
 
 
 
 
 
 
Urinalysis 
X 
 
 
Xb 
 
 
Xb 
 
 
 
Serum chemistry (mL)g 
5 
 
 
5b 
 
 
5b 
 
 
 
CBC, differential, platelets 
(mL) 
5 
 
 
5b 
 
 
5b 
 
 
 
Vital signs 
X 
X 
 
 
 
X 
 
 
 
 
Interval history 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Focused physical examinationa 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Distribute/review diary cards 
 
Xb 
 
Xb 
 
Xb 
Xb 
 
 
 
Solicited adverse events (incl. 
con. meds.) 
 
X 
X 
X 
 
X 
X 
 
 
 
Unsolicited adverse events 
(incl. con. meds.) 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
SAEs, AESIs, and SUSARs 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 


 
 
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Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
Study Visit Day  
(D) or Month (M)  
Screen 
0 
D3  
D7 
D28 
D56 
D63 
D70 
D84c  M6  (D168)d,e 
M12 (D336) 
M18 (D504) 
M24 (D672) 
(incl. con. meds.) 
Solicited injection site reaction 
adverse events (incl. con. 
meds.)  
 
X 
X 
X 
X 
X 
X 
X 
X 
 
Site of injection examination 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
TB symptom screen 
X 
 
 
 
X 
X 
 
 
X 
X 
Study vaccine administration 
 
X 
 
 
 
X 
 
 
 
 
Whole blood assay (mL) 
 
 6b  
 
 
 
 
 
6b 
 
 
Absolute blood count (mL) 
 
0.5 
0.5 
 
 
 
 
 
 
0.5 
RNA (mL) 
 
2.5 
2.5 
 
 
 
 
 
 
2.5 
PBMC for ICS (mL) 
 
8b 
 
 
 
 
 
8b 
 
 
Serum for multiplex ELISA 
(mL) 
 
3b 
3b 
 
 
 
 
 
 
 
PBMC, plasma, and serum for 
correlates of risk/protection 
(mL) 
 
16 
 
 
 
 
 
16 
 
40i 
Whole blood for evaluation of 
novel IGRA (mL)f 
7 
7 
 
 
 
 
 
 
7 
7 
PBMC for evaluation of  NTM 
exposure (mL) 
 
10 
 
 
 
 
 
 
 
 
Per visit phlebotomy volume 
(mL)h 
23 
53 
6 
10 
0 
0 
10 
32.5 
10 
53 (M6,M12)/ 
13 (M18,M24) 
Cumulative phlebotomy 
volume (mL)h 
23 
76 
82 
92 
92 
92 
102 
134.5 144.5 
276.5  
a. 
If indicated by interval history 
b. Subjects in the Safety & Immunogenicity Cohort only 
c. 
Subjects who convert from QFT-GIT(-) to (+) at Day 84 will have a final visit at 168 days ±14 days after second 
vaccination (Month 8, Study Day 224) for evaluation of SAEs, AESIs, SUSARs, and TB symptom screen 
d. The Month 6, 12, 18, and 24 study visits will be completed by subjects who did not convert from QFT-GIT (-) to 
(+) at the previous visit; the Month 24 visit will be the final visit for subjects who remain QFT-GIT(-) throughout 
the study 
e. 
Subjects who convert from QFT-GIT (-) to (+) at Month 6-24 visits will have two additional visits with repeat 
QFT-GIT testing, assessment of SAEs, AESIs, SUSARs, and TB symptom screen 84 days ±14 days and 168 days 
±14 days after conversion. Up to 10 mL of blood will be collected at each of those visits, including blood for 
absolute blood count and RNA. Subjects who initially convert to QFT-GIT(+) at Month 6 or 12 will be asked to 
have a final QFT-GIT test, blood collection for absolute blood count and RNA,  and TB symptom screen done at 
least 24 months after initial vaccination.   
f. 
May not be collected at all time points shown. 
g. Serum chemistry includes AST, ALT, alkaline phosphatase, total bilirubin, creatinine, and BUN. 
h. Blood volumes are approximate. 
i. 
Study Months 6 and 12 only. 
 
3.2 
Subject Selection  
3.2.1 
Recruitment and Informed Consent 
Subjects will be recruited from high schools or communities in the Worcester region, and 
potentially elsewhere with similar TB epidemiology, through liaison with Education Department 
officials, teachers, parent organizations, and community organizations. Various methods of 


 
 
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recruitment may be used, such as classroom information sessions, advertising, referrals, word-of-
mouth, or solicitation through subjects previously known to the clinical site.  All recruitment 
materials will be approved by the Institutional Review Board (IRB) or Independent Ethics 
Committee (IEC).  Interested subjects will be invited to participate in the informed consent and 
assent process.  Informed consent will be obtained by the use of a written consent form approved 
by the IRB or IEC and signed and dated by the parent/legal guardian at the time of consent.  
Similarly, informed assent will be obtained by the use of a written assent form approved by the 
IRB or IEC and signed and dated by the subject at the time of assent. Potential subjects will be 
interviewed to ensure that they meet all entry criteria relating to history. The clinical investigator 
or designee will conduct the consent and assent discussions on an individual basis with each 
subject and parent/legal guardian and will allow adequate time for all questions to be addressed.  
Written informed consent and assent will be obtained prior to conducting any study-related 
procedures. A copy of the signed consent and assent forms shall be given to the subject and 
parent/legal guardian prior to conducting any study-related procedures. 
 
Subjects who converted to a positive QFT-GIT at Month 6 or 12 will be approached for an 
additional QFT-GIT test, blood collection for absolute blood count and RNA, and evaluation for 
signs/symptoms of TB to be conducted at least 24 months after their initial vaccination. Informed 
consent will be obtained by the use of a written supplemental consent form approved by the IRB 
or IEC, and signed and dated by subject or by the subject’s parent/legal guardian if subject is less 
than 18 years of age. Similarly, informed assent will be obtained by the use of a written assent 
form approved by the IRB or IEC, and signed and dated by the subject if the subject is less than 
18 years of age.   
 
3.2.2 
Screening 
After informed consent is obtained, subjects will be screened to assess eligibility for the study.  
For identification purposes each subject will be assigned a unique 12-digit subject number by an 
interactive voice/web response system (IVRS/IWRS) that consists of the last two digits of the 
Aeras product number (04), the last two digits of the protocol number (40), a 2-digit site number 
assigned by Aeras, followed by a 5-digit number sequentially assigned by the system.  (For 
example, if the clinical site number is 99, the first subject to be screened for protocol C-040-404 
would receive the number 044099-00001, where all except “-00001” were pre-assigned by 
Aeras.)  This subject number will be used throughout the study.  
 
Abnormal results and findings resulting in ineligibility will be discussed with the subject, who 
will be referred for follow-up care with their healthcare provider if necessary. 
 
Eligibility for randomization will be based on the inclusion and exclusion criteria described 
below.  The investigator must document confirmation of eligibility prior to randomization. 
 
3.2.3 
Inclusion Criteria 
Subjects must meet all of the following criteria at the time of randomization: 
1. Has completed the written informed consent and assent process 
2. Is age  12 years and  17 years on Study Day 0 


 
 
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3. Agrees to stay in contact with the study site for the duration of the study, provide updated 
contact information as necessary, and has no current plans to move from the study area for 
the duration of the study 
4. For female subjects:  agrees to avoid pregnancy from 28 days prior to Study Day 0 and for 
the full duration of the study. Women physically capable of pregnancy (not sterilized and still 
menstruating or within 1 year of the last menses) in sexual relationships with men must use 
an acceptable method of avoiding pregnancy during this period.  Acceptable methods of 
avoiding pregnancy include a sterile sexual partner, sexual abstinence (not engaging in 
sexual intercourse), hormonal contraceptives (oral, injection, transdermal patch, or implant), 
vaginal ring, intrauterine device (IUD), or the combination of a condom or diaphragm with 
spermicide 
5. Has general good health, confirmed by medical history and physical examination 
6. Had BCG vaccination at least 5 years ago documented through medical history or by 
presence of healed BCG scar 
7. Tests QFT-GIT negative at screening, using the manufacturer’s recommended threshold of 
0.35 IU/mL 
 
3.2.4 
Exclusion Criteria 
Subjects must have none of the following at the time of randomization: 
1. Acute illness on Study Day 0 
2. Axillary temperature 37.5C on Study Day 0 
3. Abnormal laboratory values from the most recent blood collected prior to randomization as 
follows: 
• Laboratory evidence of hematologic disease (white blood cell count <3000/mm3 or 
>11,500/mm3; hemoglobin <0.9 times the lower limit of normal of the testing laboratory, 
by age and gender; absolute neutrophil count <1300/mm3; absolute lymphocyte count 
<1000/mm3)  
• ALT, AST, alkaline phosphatase, total bilirubin, creatinine, blood urea nitrogen (BUN) 
>1.25 times the upper limit of normal of the testing laboratory  
4. Greater than Grade 1 on the Toxicity Scale urinalysis result (with the exception of hematuria 
in a menstruating female), or urinalysis abnormality judged clinically significant by the 
investigator 
5. History or evidence of any clinically significant systemic disease, or any acute or chronic 
illness that might affect the safety, immunogenicity, or efficacy of study vaccine in the 
opinion of the investigator 
6. History of treatment for active TB disease or latent Mtb infection 
7. History or evidence, including chest X-ray, of active TB disease 
8. Shared residence with an individual receiving anti-TB treatment, or known to have 
incompletely treated culture or smear positive TB, at screening 
9. History of autoimmune disease or immunosuppression 
10. Used immunosuppressive medication within 42 days before Study Day 0 (inhaled and topical 
corticosteroids are permitted)  
11. Received immunoglobulin or blood products within 42 days before Study Day 0 


 
 
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12. Received any investigational drug therapy or investigational vaccine within 182 days before 
Study Day 0, or planned participation in any other clinical trial during the study period 
13. Received investigational TB vaccine, other than BCG, at any time prior to Study Day 0 
14. Planned administration/administration of a licensed vaccine in the period starting 28 days 
before and ending 28 days after each dose of study vaccine 
15. History or laboratory evidence of any past or present possible immunodeficiency state 
including, but not limited to, any laboratory indication of HIV-1 infection 
16. History of allergic disease or reactions, including eczema, likely to be exacerbated by any 
component of the study vaccine 
17. History of alcohol or drug abuse  
18. All female subjects:  currently pregnant or lactating/nursing; or positive urine pregnancy test 
during screening  
19. Received a tuberculin skin test (TST) within 3 months (90 days) prior to Study Day 0.   
20. Any current medical, psychiatric, occupational, or substance abuse problems that, in the 
opinion of the investigator, will make it unlikely that the subject will comply with the 
protocol 
 
3.2.5 
Screening Clinical Assessments and Laboratory Tests 
Unless noted otherwise, the window period within which all screening evaluations must be 
completed, and the results reviewed by the investigator to confirm eligibility of subjects, is 21 
days prior to and including Study Day 0.   
 
Subjects will provide a detailed medical history and undergo a physical examination. 
Demographic characteristics (date of birth, gender, and race/ethnicity) will also be collected. A 
dipstick urinalysis, urine pregnancy test (all females), QFT-GIT test, and an HIV rapid test with 
repeat confirmation of all positive tests, will be performed. Any new abnormal findings will be 
discussed with the subject and referral will be made for follow-up care if necessary. 
 
Screening laboratory tests (including serum chemistry, complete blood count, differential blood 
count, platelets, and hepatic enzymes) will be performed during the screening process. Results 
from these laboratory tests will serve as study-entry baseline values. Abnormal results and 
findings that make the subject ineligible will be discussed with the subject and the subject will be 
referred for follow-up care with their healthcare provider if necessary. All screening laboratory 
specimens will be processed according to laboratory SOPs available from the clinical 
laboratory(ies) designated for the study. Information about the laboratory(ies), including any 
instructions for performing and interpreting specific tests, will be maintained in the investigator’s 
study files. 
 
3.3 
Study Randomization 
Subjects will be randomized to the study based on a randomly-generated sequence of subject 
identification numbers (randomization schedule) managed by a validated IVRS/IWRS. The 
randomization schedule will be prepared by a statistician who will not be involved in the analysis 
of the study in order to maintain the blind of the study team.  The day of randomization for each 
subject will be Study Day 0. Randomization will be in blocks for each school or community, as 


 
 
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the epidemiology of TB transmission may vary from one school or community to another. 
Randomization will also be blocked such that 30 subjects from each vaccine group will be 
enrolled in the Safety & Immunogenicity Cohort. Enrollment for the Safety & Immunogenicity 
Cohort will be from a subset of schools or communities selected to be representative of the study 
population as a whole in terms of socio-economic status and TB risk. Subjects who discontinue 
participation will not be replaced. 
 
3.4 
Blinding 
Assignment of subjects to the BCG Vaccine SSI arm will be unblinded (i.e. subjects and clinic 
staff will be aware of study product allocation), with the exception that laboratory staff 
performing the QFT-GIT and other immunological endpoint assays will remain unaware of study 
product allocation. 
 
Assignment of subjects to the AERAS-404 and saline placebo arms will be blinded, such that 
subjects, site study staff, and laboratory staff will remain unaware of study product allocation. 
Since AERAS-404 and the saline placebo have different appearances, the syringes containing 
AERAS-404 or comparator will be masked so that the study injection administrator (the study 
team member in the clinic who will be administering the injections) can also remain blinded.  
 
The Aeras investigational product manager (and/or designee) will be unblinded in order to 
manage study vaccine inventory. The other unblinded persons on the study are the study vaccine 
manager (and designee, if appointed) and the study monitor(s) responsible for monitoring study 
vaccines. All unblinded persons must take care to not reveal individual subject treatment regimen 
assignments to any other member of the study team.   
 
The study vaccine manager (and designee) must be a designated study team member, such as the 
study pharmacist, who is not an employee of Aeras and who will have no other clinical or 
regulatory responsibilities associated with the conduct of the study during the entire study period.  
Unblinded study personnel must not participate in the evaluation of adverse events. A Delegation 
of Authority Log will be maintained by the site and will identify the individual(s) authorized to 
function as the study vaccine manager, i.e., individuals with access to study blinding 
information.   
 
Labels accompanying the syringes of prepared AERAS-404 and placebo will not indicate which 
study vaccine is in the syringe.  Identical syringes and needles will be used for preparation and 
administration of AERAS-404 and placebo.  
 
3.4.1 
Unblinding for Clinical Emergencies  
If there is an urgent clinical requirement to know a subject’s treatment assignment, the principal 
investigator will request the urgent unblinding of a subject’s treatment by completing the Subject 
Unblinding by Site form in the IWRS.  The designated study management team will be notified 
of the unblinding immediately via the IWRS system alert.   
 


 
 
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If the IWRS system has been closed or is otherwise unavailable, the investigator will make a 
written request to the vaccine manager for urgent unblinding of a participant’s treatment.  The 
request must include the subject identification number, the date, a brief justification of the 
clinical requirement for unblinding, and the investigator’s signature. The request will be kept in 
the study file. Upon receipt of proper written request, the vaccine manager or designee will 
disclose the treatment group to the investigator.  Aeras must be notified immediately of any 
clinically required break of the study blind on an Immediately Reportable Event Form. 
 
It is recommended that the principal investigator consults with the local medical monitor and 
study medical team prior to unblinding of a subject.  However, in urgent circumstances at the 
principal investigator’s discretion, the site can proceed with the unblinding of a subject without 
prior consultation with the study team. 
 
3.5 
Vaccine Administration 
On Study Day 0, subjects will receive their study injection as soon as possible after 
randomization, and after their baseline immunology blood collection and other required 
assessments.  
 
BCG 
For subjects randomized to the BCG group, a single 0.1mL dose of BCG Vaccine SSI will be 
administered intradermally in the left upper arm (deltoid region) on Study Day 0, using the 
standard Mantoux technique. BCG Vaccine SSI will be administered in an unblinded fashion. 
 
AERAS-404 and Placebo 
For subjects randomized to the blinded AERAS-404 and saline placebo groups, one injection 
will be administered intramuscularly (IM) in the left upper arm (deltoid) on Study Day 0. On 
Study Day 56, subjects who have not met any of the criteria for discontinuation of study 
injections (see Section 6.1) will receive a second injection in the right upper arm. In cases of 
short term, reversible conditions, such as acute febrile or respiratory illness or evidence of 
significant active infection, the second study injection should be deferred until the subject has 
recovered; the allowable time period for deferral of the second dose is 16 days (i.e., Study Day 
56-2/56+14 days).     
 
The study vaccine manager will send the AERAS-404 and placebo to the clinic as a unit-dose 
syringe, which will be identified with the subject identification number, date and time of dose 
preparation, and the volume prepared. A medically qualified study team member must be present 
in the clinic at the time of all study injection administrations.   
 
Before administering the injection, the study injection administrator must inspect the syringe and 
vaccine volume, checking that the syringe is identified with the correct subject identification 
number and checking the date and time the dose was prepared.    
 
The syringe will contain 0.5 mL of AERAS-404 or placebo. The study injection will be 
administered IM by the study injection administrator into the deltoid area using standard aseptic 
technique.  


 
 
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The study vaccine manager will refer to the Vaccine Management Manual (provided under 
separate cover) for detailed instructions for AERAS-404 and placebo storage and preparation. 
 
3.6 
Study Evaluations 
The 90 subjects enrolled in the Safety & Immunogenicity Cohort will have more intensive 
evaluation of safety and immunologic data. The remaining 900 subjects will be enrolled into the 
Correlates Cohort. All 990 subjects in the study will be evaluated for safety and biomarker 
outcomes, and for prevention of Mtb infection. 
 
3.6.1 
Evaluation of Mtb infection 
QFT-GIT assays will be performed at screening, on Day 84 (end of the wash-out period), on 
Month 6, and periodically thereafter until end of study, as described in Section 2.2. QFT-GIT 
assays will be performed according to site SOPs to reduce variability due to effects such as blood 
draw technique, duration of incubation, amount of blood incubated, temperature of sample 
storage, etc. The primary evaluation of Mtb infection will be QFT-GIT conversion from a 
negative to positive test, using the manufacturer’s recommended threshold of 0.35 IU/mL, at any 
time-point after Day 84 and through end of follow-up for the primary endpoint.  
 
The exploratory evaluation of Mtb infection will be defined by alternative threshold values for 
QFT-GIT conversion and reversion (Pai, 2012; van Zyl-Smit et al, 2009; Metcalfe et al, 2013); 
alternative time periods for definition of persistent QFT-GIT conversion; and development and 
validation of novel IGRAs.  
 
The South African National TB Programme (NTP) does not recommend a course of isoniazid 
preventive therapy (IPT) for HIV-uninfected adults and children older than 5 years of age with 
asymptomatic (latent) Mtb infection, due to the high risk of reinfection after completion of IPT in 
this high TB prevalence region. Subjects in this trial will be managed according to current NTP 
guidelines. However, since QFT-GIT conversion is known to be associated with increased risk of 
active TB disease, symptom screening for early diagnosis and treatment of incident TB disease 
will be a safety priority, and all subjects who are found to be QFT-GIT positive (including at 
screening) will be educated on the symptoms of TB and the benefits of early diagnosis and 
treatment. Screening for symptoms compatible with incident TB disease, including unexplained 
cough, fever, night sweats, or loss of weight for longer than two weeks duration will continue 
throughout the study. Subjects with one or more compatible symptoms will be asked to provide 
two sputum samples for smear microscopy, mycobacterial culture, and GeneXpert. Subjects who 
are sputum smear negative or unproductive may also undergo chest radiography or additional 
investigations, including repeat HIV testing, as clinically indicated. Subjects with a 
microbiological, radiological, and/or clinical diagnosis of TB disease will be referred to the NTP 
for TB treatment (directly observed, short course), which is provided free of charge by the NTP. 
For the purposes of the exploratory TB disease objective, a bacteriologically confirmed case of 
active TB disease will be defined as clinical evidence of TB with positive Mtb MGIT culture, or 
GeneXpert MTB/RIF (Cepheid, USA), from sputum or an extra-pulmonary site. A clinical case 


 
 
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of active TB disease will be defined as clinical and/or radiographic evidence of active TB disease 
that is not bacteriologically confirmed (as defined above).  
  
3.6.2 
Immunology Laboratory Evaluations 
A summary of immunologic assays to be performed on blood specimens is shown in Table 3-3. 
Staff at the clinical research site will refer to the most current version of the Specimen 
Management Manual (provided under separate cover) for further instructions and additional 
information on specimen collection and processing. All blood volumes are approximate. 
 


 
 
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Table 3-3 
Summary of Immunology Laboratory Evaluations 
Sample type 
 
Assay 
Blood 
volume 
To be  
conducted by 
Study Days (D)/ 
Study Months (M) 
Immunogenicity (Safety & Immunogenicity Cohort only, n = 90) 
Immunogenicity 
(primary) 
PBMC 
Intracellular cytokine 
staining (ICS) 
8mL (CPT) 
Aeras or South 
Africa Endpoint 
Laboratory, Cape 
Town 
D0, D70 
Immunogenicity 
(secondary) 
Whole blood in 
Sodium Heparin 
Intracellular cytokine 
staining (ICS)  
6mL (Na Hep) 
Site laboratory 
D0, D70 
Early innate signature (Safety & Immunogenicity Cohort only, n = 90) 
Serum from clotted 
blood  
Quantification of soluble 
immune mediators by 
multiplex ELISA  
3mL (clotted 
tube) 
TBD 
D0, D3 for AERAS-
404/Placebo 
 
D0, D7 for BCG 
Correlates of Protection (Safety & Immunogenicity and Correlates Cohorts, n=990) 
PBMC, plasma, 
and serum 
TBD 
Up to 16mL (2 
CPT tubes, 1 
clotted tube) 
(2mL plasma; 
2mL serum) 
TBD 
D0, D70 
Up to 40mL (4 
CPT tubes, 1 
clotted tube) 
(2mL plasma; 
2mL serum) 
TBD 
M6, M12 
Whole blood in 
PAXgenea 
Transcriptomics on RNA 
2.5mL 
(Paxgene) 
TBD 
D0, D3 for AERAS-
404/Placebo 
 
D0, D7 for BCG 
 
M6, M12, M18, 
M24, and 3 and 6 
months after QFT 
conversion for all 
groups 
Whole blood in 
Lithium Heparin 
microtainera 
Absolute blood cell subset 
counting by flow cytometry 
(deconvolution of 
transcriptomics) 
0.5mL 
Site laboratory 
D0, D3 for AERAS-
404/Placebo 
 
D0, D7 for BCG 
 
M6, M12, M18, 
M24, and 3 and 6 
months after QFT 
conversion for all 
groups 
a. 
Subjects who initially convert to QFT-GIT(+) at Month 6 or 12 will be asked to have a final blood collection for 
absolute blood count and RNA done at least 24 months after their initial vaccination. 
 


 
 
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3.6.3 
Safety Evaluations 
3.6.3.1 Pre-vaccination and Post-Vaccination Monitoring of Subjects 
Subjects will have vital signs taken prior to each study vaccination. Subjects will remain in the 
clinic under close observation for at least 30 minutes after receiving study vaccine or BCG. Vital 
signs will be repeated before subjects leave the clinic. Allergic reactions to vaccination are 
possible, therefore, appropriate drugs and medical equipment to treat acute anaphylactic 
reactions must be immediately available and a medically qualified study team member trained to 
recognize and treat anaphylaxis must be present in the clinic during the entire vaccination 
procedure and post-vaccination monitoring period. 
 
3.6.3.2 Clinical Assessments and Laboratory Tests 
Clinical assessments and laboratory tests to be performed at each visit are summarized in Tables 
3-1 and 3-2. 
 
Abnormal Clinical Assessments and Laboratory Tests 
Results from clinical laboratory tests obtained on the study must be promptly reviewed by the 
investigator (or a designee who is a medically qualified study team member) as per site policy to 
determine if abnormalities exist.  If the laboratory value is abnormal and has increased in toxicity 
grade (see Appendix C for toxicity grading scales) from pre-vaccination values, it must be 
reported as an adverse event and repeated promptly at the investigator’s discretion to 
demonstrate resolution.  Additional laboratory tests may be performed if the investigator deems 
them to be necessary to fully evaluate an adverse event. In the event that the investigator elects to 
order non-protocol-specified laboratory tests, the investigator must record the rationale for the 
tests and a determination of clinical significance of the result in the source documents. The 
investigator must keep the medical monitor informed of adverse events of clinical significance.   
 
Abnormal results and findings will be discussed with the subject, and the subject will be referred 
for follow-up with their healthcare provider if necessary. 
 


 
 
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3.6.3.3 Adverse Events 
The collection periods for adverse events are shown in Table 3-4. 
 
Table 3-4 
Adverse Event Collection Periods 
Type of Event 
Collection Period 
Unsolicited adverse events 
28 days post each vaccinationa 
Solicited adverse events 
7 days post each vaccination 
(with diary cards to be used for 7 days after each vaccination for Safety & 
Immunogenicity Cohort only) 
Solicited and unsolicited injection site 
reaction adverse events 
BCG Group:  84 days post vaccination 
AERAS-404/Placebo Groups:  28 days post each vaccinationa  
Serious adverse events, adverse events 
of special interest, and SUSARs 
Entire study period 
a. 
Adverse events will be collected through the end of the study visit window for each adverse event collection period 
(i.e., 28 days after each vaccination + 5-day window = 33 days) 
 
For this study, solicited adverse events to be collected include the following:  pain, redness and 
swelling at the site of injection; fever; myalgia; arthralgia; fatigue; headache; anorexia; hives; 
and chills. Solicited injection site reaction adverse events include (in addition to pain, redness, 
and swelling) ulceration, drainage, and axillary lymphadenopathy. Solicited adverse events of 
local injection site reactions (i.e., pain at injection site, redness at injection site, swelling at 
injection site, ulceration at the injection site, or drainage at the injection site) will be considered 
causally related to study injection (adverse reaction). Adverse events of special interest (see 
Appendix D) will be collected passively during participant safety evaluations. 
 
3.6.3.4 Concomitant Medications 
The collection of information on concomitant medications used by subjects following 
vaccination will coincide with the collection period of adverse events (solicited and unsolicited 
adverse events; and solicited and unsolicited injection site reaction adverse events; see Table 
3-4). In addition, information on all concomitant medications associated with the treatment of 
serious adverse events and adverse events of special interest will be collected throughout study 
follow up. Information on any medications being taken to treat TB disease will also be collected 
throughout the study.  
 
Concomitant medication includes prescription and non-prescription drugs or other treatments, 
and any vaccines other than the study vaccines. The name of the medication, treatment start and 
stop dates (or ‘ongoing’), route of administration, and indication must be recorded on the 
Concomitant Medications case report form (CRF). The indication recorded on the Concomitant 
Medications CRF must correspond to a medical term/diagnosis recorded on the adverse event 
(AE) CRF, or to a pre-existing condition noted in the subject’s medical history, or be noted as 
prophylaxis, e.g., dietary supplement. 
 


 
 
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3.6.4 
Subject Follow-up and Contact 
All subjects who are assigned a subject identification number and receive study vaccine will be 
followed according to the protocol unless consent is withdrawn.   
 
Subjects will be instructed to contact a study team member to report new diagnoses or new or 
worsening adverse events and to come to the study clinic if medical attention is needed, provided 
the urgency of the situation permits.  For emergencies and other unscheduled visits to a medical 
facility other than the study clinic, medical records will, to the extent possible, be obtained by the 
investigator.   
 
Study visits will be scheduled out of school hours (if applicable) whenever possible. During each 
clinic visit, subjects will be reminded to notify a study team member of the following:   
• 
The occurrence of AEs and SAEs during the respective reporting periods  
• 
Receipt of any concomitant medications during the applicable reporting period 
• 
Plans to move or if contact information changes  
• 
If the subject has decided to withdraw from the study 
• 
Change in general health status  
• 
Any other change in status that may affect the subject’s participation (e.g., plan to participate 
in another clinical trial) 
 
All deviations from protocol procedures, evaluations, and/or visits will be documented according 
to the Case Report Form Guidelines provided by Aeras Data Management or their designee.  
When possible, missed visits and procedures must be rescheduled and performed at the nearest 
possible time point to the original schedule. 
 
3.6.5 
Loss to Follow-up 
If the site’s study team members are unable to establish contact with a subject who misses a 
scheduled study visit, the clinical site must make every possible effort to re-establish contact and 
document such efforts.  If contact is re-established, then the subject will resume participation in 
the study. 
 
If contact with the subject cannot be re-established by the subject’s calculated final study visit 
date, then a determination of “lost to follow-up” can be made. 
 
4 
VACCINES  
4.1 
Supplies 
AERAS-404 
AERAS-404 is an investigational vaccine manufactured by Sanofi Pasteur (Toronto, Canada) 
and Statens Serum Institut (SSI; Copenhagen, Denmark). AERAS-404 has two components: the 
H4 antigen and the IC31 adjuvant. The reconstitution of the vaccine components, H4 antigen 
(manufactured at Sanofi Pasteur) and IC31 adjuvant (supplied by SSI), will take place at the 
study clinic. 
 


 
 
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H4 antigen is composed of a purified recombinant fusion protein. The H4 antigen is presented as 
a sterile, clear, colorless, or slightly yellow solution in single dose vials.  Each 0.5 mL of H4 
antigen solution contains 75 mcg of H4 antigen at a concentration of 150 mcg/mL. H4 antigen is 
formulated in a buffer consisting of 10 mmol/L Tris-HCl, pH 8.3. The presentation is 0.20 
mL/vial (label claim) with an actual fill volume of 0.50 mL ± 0.05 mL.  The AERAS-404 
antigen will be mixed further with adjuvant prior to injection (reconstitution). 
 
IC31 adjuvant is composed of ODN1a and KLK peptide. When thawed, the single-dose IC31 
adjuvant vials contain 0.8 mL of translucent adjuvant solution.  Each 0.8 mL of IC31 solution 
contains the following components: IC31 adjuvant at 1000 nmol KLK + 40 nmol ODN1a 
solubilized in 10 mmol/L Tris-HCl, pH 7.4, 168.75 mmol/L sodium chloride.  
 
Control Product (Placebo) 
The normal saline placebo (control) will be sourced by the study vaccine manager in sterile 
ampoules or vials.  A separate ampoule or vial will be used for each participant in order to ensure 
sterility.  
 
BCG 
BCG (BCG Vaccine SSI, powder and solvent for suspension for injection) will be supplied by 
the study site.  After reconstitution, 1 dose (0.1 mL) for adults and children aged 12 months and 
over contains Mycobacterium bovis BCG (Bacillus Calmette-Guerin), Danish strain 1331, live 
attenuated, 2-8 x 105 cfu.  
 
4.2 
Accountability 
The study vaccine manager is required to maintain accurate study vaccine accountability records.  
Instructions and forms to be completed and kept for accountability will be provided to the study 
vaccine manager.  If the study vaccine manager wishes to use site-specific accountability forms, 
these must be reviewed and approved in advance by Aeras.  Upon completion of the study, all 
study vaccine management records will be copied and the copies returned to Aeras or its 
designee.  The originals must be maintained at the clinical site with the rest of the study records. 
 
4.3 
Receipt and Storage 
Upon receipt of study vaccine supplies, the study vaccine manager must immediately inspect all 
vials for damage. AERAS-404 (H4 antigen and IC31 adjuvant) will be shipped with a continuous 
temperature-monitoring device.  Any damage or discrepancies from the packing list must be 
documented and promptly discussed with Aeras and the study monitor to determine the 
appropriate action.  
 
AERAS-404 (H4 antigen and IC31 adjuvant) must be stored at less than or equal to -15°C in a 
secured location with no access for unauthorized personnel. BCG must be stored at 2-8°C in a 
secured location with no access for unauthorized personnel. The normal saline placebo will be 
stored at room temperature in the study pharmacy. 
 


 
 
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Refer to the most recent version of the Vaccine Management Manual for detailed instructions 
regarding study vaccine storage. 
 
4.4 
Vaccine Preparation 
AERAS-404 and Control (Placebo) 
Refer to the most recent version of the Vaccine Management Manual (VMM) for detailed 
instructions regarding AERAS-404 and placebo preparation. 
 
BCG 
BCG will be prepared and administered as per the manufacturer’s recommendations.  BCG 
Vaccine SSI will be prepared by the study pharmacist from multi-dose vials dispensed according 
to the package insert using aseptic technique. Each vial of BCG Vaccine SSI will be 
reconstituted with diluted Sauton SSI as specified in the package insert. Reconstituted vaccine 
may be kept at 4-8°C for up to 4 hours. Exposure to light should be kept to a minimum. Any 
reconstituted vaccine not used within 4 hours must be discarded. 
 
4.5 
Disposal of Unused Supplies 
Aeras must provide authorization for any unused study vaccine and supplies to be destroyed.  
Unused supplies will be destroyed according to the facility’s SOPs.  Any disposal of study 
vaccine conducted at the clinical site must be documented in the study file. 
 
5 
SAFETY  
5.1 
Responsibilities for Ensuring the Safety of Trial Subjects 
The national regulatory authority, the study sponsor (Aeras), the institution through which the 
research is performed and all members of the principal investigator’s clinical team share 
responsibility for ensuring that participants in this trial are exposed to the least possible risk of 
adverse events that may result from participation in this protocol.  
 
5.1.1 
Principal Investigator 
The principal investigator has a personal responsibility to closely monitor trial subjects and an 
inherent authority to take whatever measures necessary to ensure their safety.  The principal 
investigator has the authority to terminate, suspend or require changes to a clinical trial for safety 
concerns and may delay an individual’s study vaccine administration or pause study vaccine 
administration in the whole trial if the investigator has some suspicion that the study vaccine 
might place a subject at significant risk. Where specified, the responsibilities of the principal 
investigator may be delegated to a medically qualified team member (designee). The principal 
investigator or designee determines severity and causality with respect to the study vaccine for 
each adverse event. For blinded studies the principal investigator is blinded, in which case the 
study vaccine may consist of a placebo, an active control, or the investigational product.  
 


 
 
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5.1.2 
Study Sponsor 
The sponsor (Aeras) also has an institutional responsibility to ensure subject safety.  This 
responsibility is vested in a local medical monitor and a data monitoring committee (DMC).   
 
5.1.3 
Local Medical Monitor 
The local medical monitor is the sponsor’s representative and is a physician or surgeon in their 
country of residence.  The local medical monitor reviews the safety of the product for protocols 
in a specific region and, in conjunction with the sponsor, determines expectedness of the adverse 
event.  The local medical monitor, in consultation with the sponsor, may assess the severity and 
causality for adverse events and may upgrade the degree of severity and causality determined by 
the principal investigator or designee.  The local medical monitor, like the principal investigator, 
will be blinded for the subjects in the AERAS-404 and placebo study groups.  
 
5.1.4 
Data Monitoring Committee 
The DMC will operate according to its charter.  If study vaccine administration is paused by the 
local medical monitor or the principal investigator, the DMC will be convened.  Based on its 
review and the protocol stopping rules (Section 6.2) the DMC will make recommendations in the 
DMC minutes to Aeras regarding further conduct of the study and further administration of study 
vaccine.  The DMC may review an individual SAE or it may choose to review adverse events, 
serious adverse events, solicited adverse events, and laboratory and vital signs data.  The DMC 
may unblind any amount of safety information needed to conduct their assessment.  All 
procedures associated with this review, including objectives, data handling, and elements to be 
included for review will be documented in DMC minutes. The DMC may recommend 
suspension or resumption of enrollment and study vaccine administration after review of safety 
data. However, the sponsor will make the final decision to suspend or resume study activities. 
The recommendations of the DMC, along with the sponsor’s decision, will be communicated to 
the investigators and the IRB/Ethical Committees and the national regulatory authorities. The 
sponsor or its designee agrees to abide by any directives issued by the national regulatory 
authority or the Institutional Review Board.   
 
The DMC will review safety data for the Safety & Immunogenicity cohort as described in 
Section 2.2. The DMC will also review unblinded data after sufficient QFT-GIT conversion 
endpoints have accumulated for the primary analysis of prevention of infection with Mtb, and if 
an impact on infection (positive or negative) is seen will advise the Sponsor accordingly. The 
Sponsor may then elect to extend follow up of all subjects to explore the duration of prevention 
of infection after vaccination, or to terminate the study.     
 
5.1.5 
Institutional Review Boards and Ethics Committees 
The Institutional Review Board or Ethics Committee has institutional responsibility for the safety 
of research subjects. The Institutional Review Board or Ethics Committee has the authority to 
terminate, suspend or require changes to a clinical trial.   
 


 
 
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5.1.6 
National Regulatory Authority 
Since the national regulatory authority (such as the Medicines Control Council for South Africa) 
receives all expedited safety reports it also has the authority to terminate, suspend or require 
changes to a clinical trial.   
 
5.2 
Safety Surveillance During the Study 
Subjects will be monitored and safety data collected by way of clinical interviews and 
examinations, evaluations of daily diaries conducted by study team members, and through 
reports of laboratory evaluations.  Time points and the specific data collected for each of these 
evaluations are described in Section 3 and the protocol appendices.  
 
The principal investigator and local medical monitor will review safety data on an ongoing basis 
to ensure that study pausing rules are not met.  
 
5.3 
Definition of Adverse Event 
An adverse event (AE) is any untoward medical occurrence in a patient or clinical investigation 
subject administered a pharmaceutical product and which does not necessarily have a causal 
relationship with this treatment. An adverse event can therefore be any unfavorable and 
unintended sign (including abnormal laboratory findings), symptom or disease temporally 
associated with the use of a medicinal (investigational) product, whether or not related to the 
medicinal (investigational) product.  
 
All conditions that exist prior to administration of the study vaccine (pre-existing conditions) 
will be recorded in the subject’s medical history to establish baseline.  Day-to-day fluctuations in 
pre-existing conditions that do not represent a clinically significant change in the subject’s status 
will not necessarily be reported as adverse events. 
 
Any adverse change from the subject’s baseline condition (determined from screening 
evaluations conducted to confirm study eligibility) that occurs following the administration of 
the study vaccine will be considered an adverse event.  This includes the occurrence of a new 
adverse event or the worsening of a baseline condition, whether or not considered related to the 
study vaccine.  Intermittent conditions such as headaches in adults or irritability in infants may 
be present on Study Day 0 but may represent an adverse event if the intensity or duration of the 
event is worse than usual following receipt of study vaccine.  Adverse events include but are not 
limited to: adverse changes from baseline that represent increases in toxicity grade according to 
the Toxicity Table (see protocol appendices), adverse changes in the general condition of the 
subject, signs and symptoms noted by the subject, concomitant disease with onset or increased 
severity after study vaccine administration, and changes in laboratory safety parameters 
occurring after study vaccine administration.   
 
The reporting period for all adverse events is specified in Section 3.  Adverse events will be 
reported on the Adverse Event CRF using a recognized medical term or diagnosis that accurately 
reflects the event.  Adverse event evaluations will be reviewed by the principal investigator or by 
a designated medically qualified practitioner.  Adverse event CRF pages are to be completed by 


 
 
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members of the study team designated in writing by the principal investigator.  The onset and 
resolution dates of the event and action taken in response to the event will be documented.  All 
adverse events must be followed until resolution is demonstrated.  The resolution date will be 
recorded on the CRF as the last date on which the subject experienced the adverse event.  If an 
adverse event resolution date is uncertain the principal investigator or designee should estimate 
the completion date based on medical judgment and interview of the subject. Approximate dates 
of resolution from interviews may be taken as adverse event resolution dates.  Some examples of 
estimation of adverse event resolution are:  1) an asymptomatic laboratory abnormality on one 
visit that has not been followed-up between visits but has resolved by the next visit may be 
assumed to have resolved by the midpoint of the intervisit interval; 2) A resolved adverse event 
that was treated may be assumed to have been resolved by the end of treatment.  Adverse events 
that are still present at the end of the trial should be recorded as ongoing.  Information recorded 
on the CRF must be substantiated in the source documents.  If an adverse event evolves into a 
condition that becomes “serious,” it will be designated as serious on the Adverse Event CRF and 
a Supplemental SAE Report (SAER) form will be completed.  
 
5.4 
Assessing Severity 
The safety concepts of “severity” and “seriousness” are distinct concepts (see Section 5.8).  
Severity refers to a degree of clinical manifestation.  “Seriousness” refers to defined outcomes 
from an adverse event.  A severe adverse event is not always serious and a serious adverse event 
is not always severe.  
 
For all adverse events, the investigator (or designee, who is a healthcare professional; is someone 
the investigator deems qualified to review adverse event information, to provide a medical 
evaluation of the event, and to classify the event based upon medical judgment and the severity 
categories described below) is responsible for assessing the severity of the event and the causal 
relationship of the event to the study vaccine. 
 
The severity of all adverse events, including clinical findings and abnormal laboratory values, 
will be classified as one of the following grades:  
1. Mild  
2. Moderate  
3. Severe   
 
A Toxicity Table is provided in the protocol appendices for the assessment of severity of 
specified adverse events.  The Toxicity Table Adverse Event Grades do not correlate directly 
with the classical severity grades of mild, moderate and severe.  FOR THE PURPOSES OF 
RECORDING EVENTS ON THE CRF, Toxicity Table Grade 1 events will be considered mild 
in severity, Toxicity Table Grade 2 events will be considered moderate in severity, and both 
Toxicity Table Grade 3 and 4 events will be considered as severe.  In the Toxicity Table certain 
local reactions such as erythema (redness) and swelling are graded according to size.  Laboratory 
values are graded according to level of deviation from the normal range.   
 
For adverse events not listed in the Toxicity Table determination of severity requires some level 
of interpretation as outlined below.  The degree of incapacity caused by the adverse event and 


 
 
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the level of medical intervention required for treatment may be helpful in assessing the overall 
severity of the adverse event.  
For example: 
• 
“Mild” events are generally regarded as noticeable but have no impact on normal activities; 
they may or may not require over-the-counter treatment managed by the subject.   
• 
“Moderate” events generally have some impact on an individual’s normal activities and may 
require general symptomatic medical intervention by a healthcare professional or by the 
subject.   
• 
“Severe” adverse events may be incapacitating, leading to suspension of normal daily 
activities, and would generally require more immediate medical evaluation and intervention 
by a healthcare professional.   
 
A change in severity of an adverse event will not be recorded as a new adverse event.  Only the 
highest severity level that occurs during the entire period of the adverse event will be recorded 
on the CRF with the onset and resolution dates encompassing the entire duration of the event.   
 
5.5 
Assessing Causal Relationship (Relatedness) 
For all adverse events, the investigator and the sponsor (the local medical monitor) will 
determine a causal relationship, to the study vaccine without knowledge, for the blinded portion 
of the study, of whether AERAS-404 or placebo was administered.  A number of factors will be 
considered in making this assessment, including:  1) the temporal relationship of the event to the 
administration of the study vaccine 2) whether an alternative etiology has been identified and 3) 
biological plausibility.  The investigator will use the following guidelines to assess the causal 
relationship of an adverse event to study vaccine:   
 
• 
Not Related to study vaccine (i.e., there is no evidence of a causal relationship; another 
etiology is known to have caused the adverse event.  The alternative etiology should be 
documented in the subject’s study record). 
• 
Unlikely Related to study vaccine (i.e., there is less than a reasonable possibility that the 
adverse event was caused by study vaccine). 
• 
Possible relationship to study vaccine (i.e., there is a reasonable possibility that the adverse 
event was caused by study vaccine.  There must be a plausible mechanism for the event to be 
related to study vaccine.  The evidence is inadequate to accept or reject, or favors rejection 
of, a causal relationship; an association exists between the event and the study vaccine but 
there may also be an alternative etiology, such as characteristics of the subject’s clinical 
status or underlying condition).   
• 
Probable relationship to study vaccine (i.e., it is likely that the adverse event was caused by 
administration of the study vaccine.  The evidence favors acceptance of a causal relationship; 
an association exists between the event and receipt of the study vaccine and there is a 
plausible mechanism for the event to be related to the study vaccine, and an alternative 
etiology is not apparent). 
• 
Definite relationship to study vaccine (i.e., the study vaccine is known to be the cause of the 
adverse event.  The evidence establishes a causal relationship; an association exists between 
the event and receipt of the study vaccine and there is a plausible mechanism for the event to 
be related to the study vaccine, and causes other than the study vaccine have been ruled out).  


 
 
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The principal investigator and the local medical monitor both determine causality.  It is expected 
that communication and consultation may occur in the assessment of the causality of adverse 
events.  The greatest degree of causal relationship (definite > probable > possible > unlikely 
related > not related) determined by either the investigator or local medical monitor after their 
discussions will determine the ultimate classification of the adverse event.  Definite, probable 
and possible are considered to be related.  Not related and unlikely related are considered to be 
unrelated. 
 
Every effort should be made by the investigator to determine the existence of any pre-existing 
conditions (e.g., headache in adults or rashes in infants on Study Day 0 with onset prior to study 
vaccination) that must be taken into consideration when assessing causal relationship of an 
adverse event.  Pre-existing conditions should be recorded in the CRF as baseline history and 
substantiated by appropriate source documentation.  Intermittent conditions such as headaches in 
adults or irritability in infants may not be present on Study Day 0 but may represent an adverse 
event if the intensity or duration of the event is worse than usual following study vaccine. 
 
5.6 
Definition of Adverse Reaction 
An adverse reaction is an adverse event judged to be related to study vaccine (see Section 5.3 for 
adverse event definition). 
 
Related adverse events (adverse reactions) are defined as those judged by the investigator 
or local medical monitor to be possibly, probably, or definitely related to study vaccine.  
 
5.7 
Solicited Adverse Events and Injection Site Reactions 
Solicited adverse events are events the subject is specifically asked about.  These adverse events 
are commonly observed soon after receipt of vaccines.  For this study, solicited adverse events to 
be collected include: pain, redness and swelling at the site of injection; fever; myalgia; arthralgia; 
fatigue; headache; anorexia; hives; and chills.  Solicited injection site reaction adverse events 
include (in addition to pain, redness, and swelling) ulceration, drainage, and axillary 
lymphadenopathy. Solicited adverse events of local injection site reactions (i.e., pain at injection 
site, redness at injection site, swelling at injection site, ulceration at injection site, or drainage at 
injection site) will be considered causally related to study vaccine (adverse reaction). 
 
The reporting period during which solicited adverse events will be evaluated is specified in 
Section 3.  The solicited adverse event reporting period begins with the day of vaccination. All 
subjects in the Safety & Immunogenicity cohort will be provided a diary card to record 
temperature and information regarding occurrences of these specific events for the 7 days post-
vaccination (see further details in Section 5.16).    
 
Adverse events and solicited adverse events including assessment of local injection site reactions 
will be assessed by the investigator for severity, causal relationship to the study vaccine, possible 
etiologies, and whether the event meets criteria as a serious adverse event (and therefore requires 
immediate notification to the medical monitor).   


 
 
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Presence of ulceration and/or scarring at the site of injection and axillary lymphadenopathy of 
the injection arm(s) are considered to be adverse events that are causally related to the study 
vaccine and are of special interest.  Site of injection ulceration (including presence of drainage) 
and axillary lymphadenopathy will be actively evaluated during through Day 84.  These events 
will be recorded on the Adverse Event CRF as applicable. 
 
Visible injection-site abnormalities other than expected site reactions should be photographed. 
The types of events to be photographed should be findings that are grossly abnormal or beyond 
the expected injection site reactions. These types of reactions may be reactions that appear 
rapidly or are larger or more severe than usual. The purpose of the photograph is for information 
sharing purposes with the sponsor. The types of events that should be photographed include, but 
are not limited to, allergic type reactions, such as prompt injection site redness, angioedema, and 
anaphylactic reaction. 
 
In the event that the clinical presentation meets the definition of a serious adverse event, an 
SAER form must be completed and the event reported per protocol instructions.  
  
5.8 
Adverse Events of Special Interest (AESIs) 
Adverse events of special interest (AESIs) are clinical events which are potentially immune 
mediated, and are listed in Appendix D. 
   
5.9 
Assessing "Seriousness" and Serious Adverse Events 
Seriousness refers to the outcome of an adverse event.  Seriousness is determined by both the 
principal investigator and the local medical monitor.  If either principal investigator or local 
medical monitor determines an event to be serious, it will be classified as such.  If any of the 
following outcomes are present then the adverse event is serious: 
 
• 
It results in death (i.e., the AE caused or led to the fatality).  Serious does not describe an 
event which hypothetically might have caused death if it were more severe. 
• 
It was immediately life-threatening (i.e., the AE placed the subject at immediate risk of 
dying.  It does not refer to an event which hypothetically may have led to death if it were 
more severe).  
• 
It required inpatient hospitalization or prolonged hospitalization beyond the expected length 
of stay.  Hospitalizations for scheduled treatments and elective medical/surgical procedures 
related to a pre-existing condition that did not increase in severity or frequency following 
receipt of study vaccine, are not serious by this criterion.  Hospitalization is defined as a 
hospital admission or an emergency room visit for a period greater than 24 hours.  
• 
It resulted in a persistent or significant disability/incapacity (i.e., substantial reduction of the 
subject’s ability to carry out activities of daily living). 
• 
It resulted in a congenital anomaly or birth defect (i.e., an adverse finding in a child or 
fetus of a subject exposed to the study vaccine prior to conception or during pregnancy). 
• 
Other medically important conditions that may not result in death, threaten life or require 
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considered a serious adverse event when, based on appropriate medical judgment, they may 
jeopardize the subject and require medical or surgical intervention to prevent one of the 
serious outcomes listed in these criteria (e.g., allergic bronchospasm requiring intensive 
treatment in an emergency room or at home; blood dyscrasias or convulsions that do not 
result in hospitalization, or the development of drug dependency or drug abuse). 
 
A serious adverse event is an adverse event meeting the outcome criteria for seriousness 
regardless of relationship to an administered medicinal product. 
 
5.10 
Assessing Expectedness 
Expected adverse events are adverse events consistent with the applicable product information 
provided by the sponsor (the investigator’s brochure for an investigational product).  The 
sponsor, in the person of the local medical monitor, determines expectedness.  If the assessment 
is that the adverse event is expected no further action is required.  If the local medical monitor’s 
assessment is that the adverse event is unexpected, then the event may represent a SUSAR or 
expedited SAE (see Sections 5.10 and 5.11). 
 
5.11 
Definition of Suspected Unexpected Serious Adverse Reaction (SUSAR) 
When an adverse event is judged to be related to an investigational product, such as AERAS-
404, and also is judged to be serious and unexpected, it is a SUSAR (suspected unexpected 
serious adverse reaction) and is subject to expedited reporting. 
 
5.12 
Reporting of Serious Adverse Events 
All serious adverse events, which include SUSARs, are reported to the sponsor and to the World 
Wide Safety Center for the entire study period (see protocol appendices).  SUSARs are reported 
even after the trial is over, if the sponsor, local medical monitor or principal investigator 
becomes aware of them.  The site will be provided with specific reporting procedures including 
the SAE eCRF and any supplemental reporting forms to be used.  Serious adverse events will be 
reported on the SAE eCRF using a recognized medical term or diagnosis that accurately reflects 
the event.   
 
Serious adverse events will be assessed for severity, causal relationship to the study vaccine, and 
expectedness by the investigator and the local medical monitor according to their roles (as 
described in Sections 5.1.1 and 5.1.3).  The onset and resolution dates of the event and medical 
care taken in response to the event will be documented.  If the event has not resolved by the final 
study visit, it will be documented as “ongoing” on the eCRF, however, follow-up of the SAE 
must continue until resolved or the condition has stabilized.  Information recorded on the eCRF 
must be substantiated in the source documents. 
 
The SAE eCRF for that event must be completed by the principal investigator or his/her 
designee, within one business day of the clinical site becoming aware of the event.  The SAE 
eCRF should be completed with all information known at the time. In case the eCRF cannot be 
completed, the Supplemental SAE Report (paper form) should be completed by the principal 


 
 
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investigator or his/her designee, and scanned and emailed, or faxed to the World Wide Safety 
Center and to the local medical monitor.    
 
Fatal or life-threatening serious adverse events that the investigator suspects are related to the 
study vaccine should be telephoned to the local medical monitor immediately upon the 
investigator’s awareness of the event.  If the local medical monitor is required by the protocol or 
chooses to suspend enrollment s/he shall immediately create a written memorandum for record to 
the study file and telephonically notify the sponsor of this act. 
 
Contact information for all safety personnel are contained in the Team Contact List which will 
be stored on site in the Site Regulatory Binder and maintained by the study sponsor.   
 
Investigators must not wait to collect additional information to fully document the event before 
notifying the local medical monitor and World Wide Safety Center of a serious adverse event.  
The initial notification should include the following (at minimum):  
 
• 
Protocol number and name and contact number of the investigator 
• 
Subject ID number (and initials and date of birth, if available) 
• 
Date subject received study vaccine     
• 
Serious adverse event(s) and date of event onset 
• 
Current status of subject  
 
Aeras has authorized the World Wide Safety Center to execute its responsibilities for expedited 
safety report submission to the appropriate regulatory authorities within specific time periods of 
being notified of the event (within 7 or 15 calendar days depending the character of the SUSAR); 
therefore, it is important that the investigator submit additional information requested as soon as 
it becomes available.   
 
Aeras will notify the DMC and the manufacturer of AERAS-404 of all SUSARs within 3 
working days of becoming aware of an event and will provide all follow-up information in a 
timely manner. 
 
5.13 
Other Events Requiring Immediate Reporting 
The investigator must report the following events by scanning and emailing, or faxing the 
appropriate form to the local medical monitor within 24 hours of becoming aware of the event:   
 
• 
Withdrawal of consent during the study for medical reasons (Immediately Reportable Event 
Form) 
• 
Emergency unblinding (Immediately Reportable Event Form) 
• 
Protocol violation affecting the safety of a subject or involving the vaccination process 
(Immediately Reportable Event Form) 
• 
Adverse event thought to be an allergic reaction to the study vaccine (Immediately 
Reportable Event Form, unless event meets SAE criteria)  
• 
Any event that, in the opinion of the investigator, precludes further administration of the 
study vaccine (Immediately Reportable Event Form, unless meets SAE criteria)  
• 
Pregnancy (Immediately Reportable Event Form, and Pregnancy Notification Form) 


 
 
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5.14 
Adverse Event Treatment, Follow-up, and Outcome 
Treatment of any adverse events will be determined by the investigator using his/her best 
medical judgment and according to current clinical practice guidelines.  All applied measures as 
well as follow-up will be recorded in the appropriate CRF.   
 
Adverse events will be considered resolved when the condition returns to normal or returns to 
the subject’s baseline status as established on Study Day 0, or when the condition has stabilized 
with the expectation that it will remain chronic. 
 
The investigator will continue follow-up on adverse events, including laboratory abnormalities 
and solicited adverse events, until the event has resolved, is otherwise satisfactorily explained, or 
the subject completes the study. 
  
Follow-up for serious adverse events must continue until resolution and the outcome reported to 
Aeras, even if this extends beyond the serious adverse event reporting period (i.e., after the final 
study visit).  For analysis purposes, the outcome for serious adverse events will be determined on 
the final study visit. 
 
Outcome of all adverse events will be classified as one of the following: 
• 
Resolved  
• 
Resolved with sequelae 
• 
Ongoing 
• 
Death 
 
If at any time after completion of the serious adverse event reporting period (the final study visit) 
the investigator becomes aware of a serious adverse event that is suspected by the investigator to 
be related to the study vaccine, the event must be reported to Aeras. 
 
5.15 
Follow-up of Subjects Who Become Pregnant  
If a subject becomes pregnant during the study, she should be encouraged to continue in the 
study for safety follow-up.  Follow-up should continue for pregnancy outcome including 
premature terminations, and data are to be included in the safety reports.  
 
The investigator must notify the local medical monitor of the pregnancy immediately (even if 
already known to have resulted in spontaneous or elective abortion) by emailing the scanned 
copy or faxing the Pregnancy Notification Form to the medical monitor.  At a minimum, the 
estimated date of conception, the estimated due date, and the date the subject received the study 
vaccine should be provided.   
 
If a subject becomes pregnant, she will not have any interventions done as normally mandated by 
the protocol.  The subject will undergo all other evaluations according to the Summary 
Schedule(s) of Evaluations.      
 


 
 
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The health status of the mother and child, the date of delivery, and the child’s sex, birth weight 
and multiparity should be reported to the safety monitor after delivery, using a Pregnancy 
Notification Form.  If delivery occurs before the final study visit, the subject should continue to 
be followed for SAEs through the final study visit unless withdrawal of consent has occurred.  If 
delivery occurs after the final study visit, the investigator should attempt to maintain contact with 
the subject to obtain information after delivery.       
 
Pregnancy will not be recorded as an adverse event.  However, pregnancy outcomes will be 
recorded in the World Wide Safety Database.  If the pregnancy results in a miscarriage or a 
planned termination, the event (spontaneous abortion or elective abortion) will be reported as an 
adverse event or serious adverse event per the investigator's judgment (e.g., if it was a medically 
important or life-threatening event that meets the definition of a serious adverse event).  
 
A congenital anomaly or birth defect (i.e., an adverse finding in a child or fetus of a subject 
exposed to the study vaccine before conception or during pregnancy) must be reported as a 
serious adverse event. 
 
If it is determined after completion of the study that a subject became pregnant during the study,  
the subject should notify the investigator.  The pregnancy must be reported to the local medical 
monitor and the status of the mother and child after delivery will be obtained and reported, when 
possible.     
 
5.16 
Subject Diary and Daily Temperature Monitoring 
Subjects in the Safety & Immunogenicity Cohort will receive, and be instructed in, the operation 
of a daily adverse event diary and a digital thermometer to be used during the specified post-
vaccination diary period after vaccine administration.  The daily adverse event diary is a tool to 
help aid the principal investigator and/or designee to engage in a conversation with the subject 
about any AEs that may have occurred between visits.  During scheduled visits through the 
specified diary period after study vaccine administration the daily diary will be collected and 
reviewed by the principal investigator (or designee) at which time any clinical details required 
for complete understanding of the information recorded will be obtained.  If possible, diaries not 
brought to the scheduled visit should be obtained before adverse event assessment is performed 
and events discussed with the principal investigator and/or designee.  Adverse events obtained 
from the diary, as determined by the principal investigator or designee, will not be directly 
recorded onto case report forms. 
 
Any entry recorded by the subject on the diary card that differs from the opinion of the 
investigator’s evaluation of the event (e.g., the severity level of an event is changed after 
interviewing the subject) must be explained by notation in source documentation.   
 


 
 
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6 
PAUSING AND STOPPING RULES  
These rules govern the pausing and stopping of study vaccine administration at any time during 
the study such as between doses (for multiple dose studies) for an individual, between 
individuals within a single dose group, and between dose groups. 
 
6.1 
Rules for Discontinuing Study Injections in an Individual Participant 
Administration of additional study injections will be discontinued for an individual participant if 
he/she has any of the following: 
 
• An objective clinical or laboratory parameter change which meets Grade 3 or Grade 4 
severity, as defined in the protocol toxicity table, AND is judged to be possibly, probably, or 
definitely related to study injection* 
• Adverse event thought to be an allergic reaction to the study injection, including anaphylaxis 
or bronchospasm  
• Any extensive rash (>40% body surface) on the thorax, abdomen, or limbs, including but not 
limited to urticaria, generalized petechiae, or erythema multiforme judged to be possibly, 
probably, or definitely related to study injection 
• Development of active TB 
• Development of autoimmune disease or immunosuppression, or an adverse event of special 
interest 
• Receipt of investigational drug therapy or investigational vaccine (other than study injections 
received as part of this study) 
• Any event that in the opinion of the principal investigator precludes administration of any 
further study injections 
• Pregnancy 
 
*Excluding local injection site redness, induration, or ulceration that follow the normal expected 
course of BCG vaccination. 
 
Subjects who are withdrawn from vaccination will be followed for a minimum of 6 months after 
the last study vaccination for safety. 
 
6.2 
Rules for Suspension of the Entire Study 
The following rules will trigger pausing by the principal investigator, local medical monitor, or 
the sponsor of further enrollment and study vaccine administration, and DMC review of 
unblinded safety data: 
 
1. One or more SAE(s) judged POSSIBLY, PROBABLY or DEFINITELY related to 
vaccination occurs 
OR 
2. Anaphylaxis or bronchospasm within 4 hours of injection, indicative of an immediate 
hypersensitivity reaction to the study injection 
OR 


 
 
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3. It is determined that a SUSAR occurred 
OR 
4. > 15% of subjects in the safety cohort experience a Grade 3 or higher event judged 
POSSIBLY, PROBABLY or DEFINITELY related to study vaccine*, excluding local 
injection site reactions that decrease to < Grade 3 within 24 hours 
OR 
5. an adverse event pattern of concern occurred  
 
*Excluding local injection site redness, induration, or ulceration that follow the normal expected 
course of BCG vaccination. 
 
If the principal investigator, local medical monitor, or sponsor pauses administration of study 
vaccine in the study, the decision will be recorded in a memorandum to the study file and will 
trigger DMC review.  If a recommendation to resume study enrollment and study vaccine 
administration is made, the DMC will record their judgment in a memorandum to the study file 
and notify Aeras.  The DMC memorandum will be forwarded to the medical monitors and 
principal investigators.  The DMC may recommend resumption of enrollment with changes to 
the protocol if it judges that such changes will eliminate or greatly reduce the safety risks 
specified in the stopping rules. However, the final decision to resume study activities or amend 
the protocol will be made by Aeras. The clinical site will be allowed to resume activities upon 
receipt of written notification from Aeras. 
 
7 
STATISTICAL CONSIDERATIONS  
The planned statistical analyses for this study are outlined below. A detailed statistical analysis 
plan will be created and finalized prior to database lock and preparation of any unblinded 
preliminary data review and for preparation of the final study report (see Section 7.7). 
 
7.1 
Subject Populations 
The safety population will consist of all randomized subjects who received at least one dose of 
BCG or study vaccine (AERAS-404 or placebo).     
 
The per-protocol population for efficacy analyses will consist of all randomized subjects who 
received all scheduled doses of BCG, AERAS-404, or placebo and had no major protocol 
deviations (to be described in the statistical analysis plan).  The intent-to-treat population for 
efficacy and immunogenicity analyses will consist of all randomized subjects. 
 
7.2 
Demographics and Protocol Compliance 
Demographic parameters (age, gender, and race/ethnicity) and other baseline characteristics will 
be summarized by treatment group for all subjects in the safety population. 
 
Listings of randomized subjects who missed any dose of BCG, AERAS-404, or placebo and 
number (percentage) of subjects with protocol deviations (to be defined in the statistical analysis 
plan) will be presented by treatment group. 


 
 
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7.3 
Prevention of Infection (QFT-GIT Conversion) Analyses 
The primary Mtb infection endpoint will be QFT-GIT conversion from a negative to positive 
test, using the manufacturer’s recommended threshold of 0.35 IU/mL, at any time-point after 
Day 84 and through end of follow-up for the primary endpoint (see below). The frequency of 
QFT-GIT negativity/positivity will be summarized for each treatment group at screening, on Day 
84, on Month 6, and for all subsequent assessments. The log-rank statistic will be used to test the 
null hypothesis of no difference in the rates of Mtb infection over the follow-up period between 
each of the AERAS-404 and BCG groups compared to the placebo group. The log-rank test 
statistic compares estimates of the hazard functions of the vaccine groups at each observed event 
time. It is constructed by computing the observed and expected number of events in the groups at 
each observed event time and then summing these to obtain an overall summary across all time 
points where there is an event. As subjects convert their QFT-GIT tests over time (experience a 
primary endpoint) or are lost to follow up the number of subjects at risk over time will change. 
As this is a proof of concept study, these two log-rank tests (AERAS-404 compared to placebo 
and BCG revaccination compared to placebo) will be performed with a 1-sided Type 1 error of 
0.1. There will be no adjustment for multiplicity to control the Type 1 error rate over the two 
tests. The rationale for not performing this adjustment is related to the fact that the tests pertain 
to efficacy assessments of two unrelated vaccines. If these two evaluations were performed in 
two separate trials then no adjustment for multiplicity would be made. Thus, one should not be 
compelled to adjust for this multiplicity simply because the two evaluations are performed 
administratively in a single trial.  
 
The secondary Mtb infection endpoint will be the combination of QFT-GIT conversion from a 
negative to positive test, using the manufacturer’s recommended threshold of 0.35 IU/mL, at any 
time-point after Day 84 and through end of follow-up for the primary endpoint, AND persisting 
without QFT-GIT reversion from a positive to a negative test through 6 months after QFT-GIT 
conversion. The methods used for the analysis of this endpoint will be the same as that for the 
primary Mtb infection endpoint, but the number of endpoints will be smaller as subjects who 
experience primary QFT-GIT conversion but then have a reversion to negative at one or both of 
the repeat tests will not be counted. 
 
As one possible mechanism of action of vaccination is to allow initial infection (QFT-GIT 
conversion) with subsequent clearance (and QFT-GIT reversion from positive to negative), rates 
of reversion (overall and as a proportion of those who initially converted) will be presented by 
treatment group as an exploratory analysis. Rates of conversion and reversion using alternative 
QFT-GIT threshold values will also be explored by treatment group.  
 
7.4 
Immunogenicity and Other Immunology Analyses 
All immunogenicity analyses will be based on subjects who received at least one dose of study 
vaccine (AERAS-404, BCG, or placebo). Immunogenicity will be summarized for all time points 
as collected and as available for the Safety & Immunogenicity Cohort, or for all subjects. No 
imputation for missing data will be performed. Data will be transformed as appropriate prior to 
analysis.   


 
 
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Additional analyses may be performed based on subjects who complete all scheduled 
vaccinations with study vaccine (per-protocol population). 
 
7.4.1 
Immune Response Determined by Intracellular Cytokine Staining Assay  
The primary variables of interest for preliminary assessment of immune response to vaccine will 
be the percentage of CD4+ and CD8+ T cells that express IFN-γ, TNF, IL-2, IL-17, IL-22, 
CD107a, and/or CD154 alone or in combination in response to stimulation with peptide pools 
representing the entire amino acid sequence of the mycobacterial antigens Ag85B and TB10.4, 
and BCG antigens.  Due to the high background observed with CD107a, cells expressing 
CD107a in the absence of any other functional response will be ignored.  Response will be 
measured by flow cytometry in the intracellular cytokine staining (ICS) assay.  
 
Median DMSO-subtracted cytokine responses and associated 95% confidence intervals (CI) or 
other descriptive statistics as appropriate will be used to summarize percentage T cell responses.  
Summaries of T cell response will be presented by T cell type (CD4 and CD8) and by 
stimulation antigen.  Summaries will include immune response at all available pre- and post-
vaccination immunology time points, and change from pre-vaccination to post-vaccination time 
points.  
 
Whole blood ICS will be conducted as a secondary immunologic endpoint, and will measure the 
frequencies and patterns of CD4+ and CD8+ T cells expressing Th1 and Th17 cytokines 
following stimulation of whole blood with peptide pools representing the entire amino acid 
sequence of the mycobacterial antigens Ag85B and TB10.4, as well as viable BCG from the 
vaccine vial. Stained cells will be acquired on a LSR Fortessa or LSRII flow cytometer. Analysis 
of these results will be performed as outlined above. 
 
7.4.2 
Additional Exploratory Analyses 
Analytic approaches for the exploratory endpoints below will be informed by best practice and 
the most recent advances in biomarker discovery and systems biology. 
 
7.4.2.1 RNA Transcriptomics and Whole Blood Absolute Blood Cell Counts 
RNA samples will be analyzed by microarray or sequencing to measure the acute inflammatory 
response induced by vaccination and thereby determine if immunogenicity can be predicted by a 
single, early measurement, and to understand how this acute response influences the character 
and function of the adaptive response. Samples collected after QFT conversion will be used to 
explore a correlate of risk for developing TB disease which has been identified by SATVI, or to 
explore other correlates as informed by recent data. Because analysis of such data is sensitive to 
changes in blood leukocyte subsets, it is important to include a measure of blood cell subsets by 
multiparameter flow cytometry, thus absolute blood cell counts will be used to deconvolute RNA 
transcriptomic results. 
 


 
 
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7.4.2.2 Quantification of Soluble Immune Mediators by Multiplex ELISA  
Serum will be collected and analyzed for soluble immune mediators, informed by results of the 
transcriptomic analyses. 
 
7.4.2.3 Correlates of Risk/Correlates of Protection 
PBMCs, plasma, and RNA samples will be collected and stored at time points likely to show 
peak responses for cellular, humoral, and transcriptomic responses respectively, and at 6 and 12 
months (when Mtb infection will be assessed), for all subjects. If an effect of vaccination is seen 
on infection with Mtb, immunologic assays will be conducted to search for correlates of risk or 
protection. Functional assays such as mycobacterial growth inhibition may also be conducted.  
 
7.4.2.4 Novel IGRAs 
Samples obtained during this study may be used to evaluate IGRA tests using antigens other than 
those in the QFT-GIT. Such assays would be useful to test vaccines containing ESAT-6 (one of 
the stimulating antigens in QFT-GIT) in the prevention of Mtb infection model. IGRAs using 
antigens from non-tuberculous mycobacteria (NTMs) may also be evaluated, as NTMs may be a 
cause of reduced specificity of the QFT-GIT, and may also interfere with demonstration of BCG 
efficacy (Wilson et al, 1995) through increased protection in control groups. 
 
7.5 
Safety Analyses 
Safety analyses will be performed using the safety population as defined in Section 7.1.  Count 
(percentage) summaries will be presented by treatment group for all subjects in the safety 
population. 
 
7.5.1 
Adverse Events 
The safety profile of BCG revaccination, AERAS-404 and saline placebo will be described by 
treatment group.  The primary variable for evaluation of the safety profile will be the number and 
percentage of unsolicited and solicited adverse events recorded at all available post-vaccination 
time points.  For all presentations of adverse events, additional summaries based on reporting 
period of adverse events following each study vaccination may also be presented.   
 
The number (percentage) of subjects with adverse events will be summarized by MedDRA 
system organ class (SOC) and preferred term (PT).  Additional summaries will present the 
number (percentage) of subjects with adverse events by severity and by relationship to study 
vaccine; each subject will be counted once per preferred term at the greatest severity or most 
related state recorded for that term.   
 
Separate summaries of the number (percentage) of subjects with solicited adverse events will 
also be presented.  Solicited adverse events will also be summarized by severity and relationship 
to study vaccine; each subject will be counted once per preferred term at the greatest severity or 
most related state recorded for that term.   
 


 
 
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Listings will be provided for subjects with serious adverse events and adverse events of special 
interest. 
 
Listings will be provided for subjects who have discontinued prematurely due to an adverse 
event.   
 
The number (percentage) of subjects with post-vaccination clinical laboratory values or vital sign 
values recorded as newly abnormal following study vaccination and meeting toxicity mild 
criteria (Grade 1) or above as specified in the Toxicity Table (Appendix C) will be tabulated at 
each post-vaccination time point and overall.  Clinical laboratory and vital sign abnormalities 
will also be reported as adverse events and will be included in the summary of adverse events. 
 
7.5.2 
Clinical Laboratory and Vital Sign Parameters 
For each clinical laboratory parameter and vital sign parameter prespecified in the protocol, 
summary statistics for continuous parameters will be presented by treatment regimen for all pre- 
and post-vaccination assessments and for change from pre-vaccination to post-vaccination 
assessments. 
 
7.6 
Sample Size Considerations 
Sample size for the trial is based on the power to detect a reduction in the rate of infection with 
Mtb, as determined by QFT-GIT conversion. As a proof of concept study looking for evidence of 
biologic impact, the trial is designed to distinguish a QFT-GIT conversion rate reduction of 50% 
compared to placebo for each vaccine (AERAS-404 and BCG) with power of 80% and a type 1 
error rate of 10% (1-sided).  
 
Trial design calculations are performed by computer simulation. The simulations estimate the 
statistical operating characteristic of a conditional binomial test which, due to relative rarity of 
the endpoint, provides an excellent approximation to the operating characteristic of the log-rank 
tests which will be used in the analyses of trial data. The following assumptions, based on 
preliminary data from SATVI (Mahomed et al, 2011b) were used in these simulations: 
 
- Primary Mtb infection endpoint rate = 10% per year 
- Rate of loss to follow up = 7% per year 
- Average rate of enrollment = 20 subjects per week 
- Maximum duration of follow up to primary conversion = 24 months with 6-monthly study 
visits 
 
Under these assumptions, a total sample size of 990 is expected to provide 64 primary QFT-GIT 
conversion endpoints approximately 21 months after the first subject is enrolled. Follow up will 
continue until 64 endpoints are accrued and a median follow up duration of 15 months for 
individual subjects have both occurred. A 10% rate of QFT-GIT conversions per year is 
conservative (SATVI data have shown an average 17% conversion rate in adolescents); however, 
continuing to follow subjects until the required number of endpoints is accrued ensures adequate 
power for the primary endpoint.  


 
 
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Sustained Mtb infection, as measured by persistence of QFT-GIT conversion, is an important 
secondary endpoint, as clearance of initial infection could be a mechanism of action of a vaccine.  
In addition, a persistently positive result is less likely to be the result of assay technical errors. 
Preliminary data from SATVI has shown an annual reversion rate of 20% over one year 
(Mahomed et al, 2011b). Based on this rate, the power to distinguish a 50% reduction in the rate 
of persistent QFT-GIT conversion is estimated to be 70% (the power is lower with the smaller 
number of endpoints expected). The difference in the rate of reversion among those who initially 
convert will also be examined. If the rate of reversion in the placebo group is 20% and if the 50% 
reduction in the rate of persistent QFT-GIT conversion in the vaccine group is due solely to an 
increase in the rate of reversions, then the power to detect such a vaccine effect on reversion rate 
is > 90%. If some of the vaccine effect is in the prevention of initial conversion and some is in an 
increase in reversions then the secondary endpoint of persistent QFT-GIT conversion is more 
sensitive to detect impact of vaccine. 
 
If no SAEs are observed among 330 subjects in each vaccine group (AERAS-404, BCG, and 
placebo), the exact upper 95% confidence bound on the rate of SAE occurrence would be 1%. In 
the safety cohort, a sample size of 30 in each vaccine group provides 90% power to detect at 
least one AE occurring at a frequency of 7% in each group. 
 
7.7 
Plan for Statistical Summaries and Analyses 
7.7.1 
Primary Data Review 
After 64 primary QFT-GIT conversion endpoints and a median of 15 months follow up have 
both occurred a primary unblinded analysis of QFT-GIT conversion data will be conducted by an 
independent statistical group and results will be reviewed by the DMC. If an effect on QFT-GIT 
conversion is seen, follow up for the entire study cohort may be extended. Study investigators 
will remain blinded to treatment assignments for the duration of study follow up.    
 
7.7.2 
Final Study Report 
The final study report will include all available safety data, immunogenicity data, clinical 
assessments, and concomitant medications through the final study visit.  The database will be 
locked prior to unblinding and preparation of the final study report when all of the above data 
have been entered, reviewed, and all queries related to the data have been addressed.   
 
Modifications or additions to the analyses described above will be included in the relevant 
statistical analysis plan(s).  Any decisions to deviate from the planned analyses described in the 
protocol and in the statistical analysis plan will be described in detail in the final study report. 
 
7.8 
Computer Methods 
Statistical analyses will be performed using SAS® version 9.1 or later under a Windows 
operating system. 
 


 
 
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8 
DATA COLLECTION, MONITORING, AND RECORD RETENTION  
For the purpose of monitoring and auditing the study, source documentation will consist of 
existing medical records and/or study records developed and maintained by the investigator.    
 
Data recorded on source documents will be transcribed onto case report forms (CRFs) provided 
by Aeras or entered using electronic case report forms (eCRFs) using an Electronic Data Capture 
(EDC) system provided and approved by Aeras.     
 
The study will be monitored regularly by Aeras or its designee throughout the study period.  For 
studies of unapproved investigational products, all study records (source documents, signed 
informed consent forms, copies of CRFs, IRB/IEC correspondence and approval letters, study 
vaccine management records) will be kept secured for a minimum of 2 years following the 
marketing of the investigational product or for 2 years after the discontinuation of the IND (or 
CTA, etc.).  Records will be kept for longer if required by local regulations.  The investigator 
will ensure that study records are not disposed of or removed from the clinical site without prior 
notification and approval from Aeras or its designee. 
 
9 
HUMAN SUBJECTS  
9.1 
Ethics and Regulatory Considerations 
The study will be conducted according to the ethical principles set forth in the Declaration of 
Helsinki, ICH-GCP, Protection of Human Subjects (21 CFR 50), Institutional Review Boards 
(21 CFR 56), Obligations of Clinical Investigators (21 CFR 312), South African Good Clinical 
Practice Guidelines, and other local regulatory requirements.  
 
Adolescents younger than 18 years of age require the consent of a parent or legal guardian, in 
addition to their own assent, for participation in clinical research in South Africa. Therefore, 
written informed assent will be obtained from each subject, and written informed consent will be 
obtained from the parent or legal guardian of each subject, prior to any protocol-specified 
procedures being conducted. Subjects who turn 18 during the course of the study will be re-
consented as adults at the next scheduled study visit. 
 
The protocol and informed assent and consent forms will be reviewed and approved by the IRB 
or IEC of the clinical site prior to any protocol-specified procedures being conducted.  The 
investigator will inform the IRB/IEC as to the progress of the study on a regular basis, or at 
minimum, once a year.   
 
 
There are potential known and unknown risks associated with both BCG revaccination and 
AERAS-404.  
 
No AERAS-404 vaccine-related serious adverse events have occurred and AERAS-404 has 
generally been well tolerated at all doses and regimens evaluated. Injection site reactions (pain, 
swelling, and erythema) have been mild or moderate. Reactions have occurred at the site of 


 
 
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tuberculin skin tests (TST) and BCG vaccination sites as described in the IB. Systemic adverse 
reactions include mild to moderate fatigue, myalgia, headache, arthralgia, and mild to severe 
pyrexia. Asymptomatic transient isolated proteinuria was seen in subjects receiving the vaccine 
and placebo in all four trials but was more frequent in South African adults, including at baseline 
(prior to vaccination). Proteinuria did not recur with revaccination and this finding is not 
considered to be clinically significant. One subject had a reactivation of Graves’ disease; and one 
subject was diagnosed with celiac disease at the end of the study; neither event was considered 
related to study vaccine.  
 
There is considerable experience with BCG vaccine, which is licensed in South Africa for 
prevention of TB in children and adults. BCG revaccination has also been extensively studied in 
adolescents and young adults, as discussed in Section 1.6. Injection site reactions are common 
and typically include redness, induration, and a superficial ulcer which may last 2-3 months. 
Regional lymphadenopathy is also common. Complications such as extensive local ulceration, 
local subcutaneous abscesses, suppurative lymphadenitis, or severe systemic complications are 
very rare and are not expected in the healthy, HIV-uninfected, Mtb uninfected adolescents to be 
recruited in this trial. 
 
Based on previous data from baseline QFT-GIT negative adolescents at SATVI, we expect the 
overall incidence of active TB disease in the study population to approximate 0.2% (estimate 2 
cases per year of follow-up) (Mahomed et al, 2011b). Those adolescents who convert to a 
positive QFT-GIT during follow-up have increased risk of incident TB disease (approximately 
1.4% per year), compared to adolescents who remain QFT-GIT negative (Machingaidze et al, 
2012). Screening for symptoms of incident TB disease will be a safety priority, in order to ensure 
early diagnosis and treatment. In countries with very high TB prevalence, such as South Africa, a 
course of isoniazid (INH) preventive therapy (IPT) for HIV-uninfected persons with Mtb 
infection is not recommended by the South African NTP, due to the high risk of multiple 
exposures and reinfection. A randomized controlled trial of community-wide INH prophylaxis 
conducted in South Africa among gold miners at high risk of TB disease showed that short-term 
TB incidence was reduced by 63% compared to the control group. However, the effect of IPT 
wore off rapidly and no benefit was observed beyond 9 months (Aurum Institute, 2012). Even 
among HIV infected persons with latent TB, a course of IPT provides only short-term protection 
that is lost within the first year of treatment (Johnson et al, 2001). 
 
In order to minimize potential risk to subjects related to study vaccination, the DMC will review 
safety data from the initial Safety & Immunogenicity cohort and may make recommendations on 
additional measures to ensure the safety of human subjects if any safety concerns arise. 
 
Other potential risks include the risks associated with phlebotomy. The blood volume that will be 
collected at each visit, and the total blood volume that will be collected over the course of the 
study, are not deemed physiologically significant in adolescents aged 12 years and older and are 
unlikely to be associated with adverse effects in the healthy adolescents to be recruited. 
 
HIV testing will be performed with appropriate pre- and post-test counseling. Adolescents in this 
community are sensitized to HIV and TB as public health problems. Ongoing epidemiological 


 
 
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studies and clinical trials of TB vaccines at the SATVI site have not found HIV testing to be a 
barrier to participation in this community. 
 
Results of investigations, including HIV test and pregnancy test results, will only be divulged to 
the parent or legal/guardian with the subject’s permission. Clinical information will not be 
released to other parties without written permission from the subject, except as necessary for 
monitoring or auditing of the study by Aeras, or its designee or applicable regulatory authorities. 
To maintain confidentiality, subject identification numbers will be used to identify the subject’s 
laboratory specimens, source documents, CRF, study reports, etc.  All study records will be 
maintained in a secured location.   
 
Subjects will be compensated for their inconvenience and additional costs due to attendance at 
study visits in the amount of ZAR 150 per study visit. 
 
After the study has been unblinded, the subject should be informed which study intervention the 
subject received. If AERAS-404 or BCG revaccination is found to be effective in prevention of 
Mtb infection, subjects allocated to the placebo arm may be offered AERAS-404 or BCG 
revaccination if QFT-GIT is negative at the end of the study. The decision of whether or not to 
offer AERAS-404 or BCG revaccination will be made by the study sponsor in conjunction with 
the IEC. 
 
9.2 
Institutional Review Board or Independent Ethics Committee 
All the documents the IRB/IEC may need to fulfill its responsibilities, such as the protocol, 
protocol amendments, information concerning subject recruitment, payment or compensation 
procedures, etc., will be submitted to the IRB/IEC by the investigator.  The IRB’s/IEC’s written, 
unconditional approval of the study protocol and the informed consent form will be in the 
possession of the investigator/clinical site staff prior to the conduct of any protocol-specified 
procedures.  
 
Modifications to the protocol may not be implemented without prior written IRB/IEC approval 
except when necessary to eliminate immediate hazards to the subjects or when the modification 
involves only logistical or administrative aspects of the study.  Such logistical or administrative 
modifications will be submitted to the IRB/IEC in writing by the investigator, and a copy of the 
correspondence to verify the submission will be maintained.   
 
The investigator must inform the IRB/IEC of modifications to the informed assent and consent 
forms or any other documents previously submitted for review/approval, of any new information 
that may adversely affect the safety of the subjects or the conduct of the study, provide an annual 
update and/or request for re-approval, and advise the IRB/IEC when the study has been 
completed.  
 
Any documents or forms to be provided to the subject or parent/legal guardian (e.g., information 
cards, form letters from the investigator) and all forms of study advertising (flyers, brochures, 
print advertisements, radio or television scripts, etc.) must be approved by Aeras or its designee 


 
 
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prior to the clinical site submitting them to the IRB/IEC.  Approval from the IRB/IEC must be 
obtained prior to providing the documents or forms to the subject or parent/legal guardian. 
 
9.3 
Informed Consent and Assent 
The principles of informed consent in the current edition of the Declaration of Helsinki and ICH-
GCP/21 CFR 50.25 should be implemented prior to any protocol-specified procedures being 
conducted.  Informed consent by the parent/legal guardian will be documented in writing on a 
consent form approved by the IRB/IEC. Similarly, subject informed assent will be documented 
in writing on an assent form approved by the IRB/IEC.  
 
The informed consent and assent processes will be conducted in a private space, whether at the 
study clinic, at home, or at school, to maintain confidentiality. Consent and assent processes will 
be conducted in the subject’s language of choice. All relevant information should be provided in 
both oral and written form in a way that is understandable to the subject and parent/legal 
guardian. Ample time and opportunity must be given for the subject and parent/legal guardian to 
inquire about details of the study.  The written consent and assent documents will embody the 
elements of informed consent as described in the Declaration of Helsinki and will also comply 
with local regulations (e.g., South African GCP).    
 
The investigator or the investigator’s qualified designee will explain the nature of the study and 
inform the subject and parent/legal guardian that participation is voluntary and that the subject 
can leave the study at any time, without penalty or loss of benefits to which they are otherwise 
entitled.  The subject and parent/legal guardian must be informed about the study’s purpose 
including why the subject was selected to participate, study goals, expected benefits and risks, 
potential risks, and that some potential risks are unforeseeable.  The subject and parent/legal 
guardian must be provided with a description of the procedures and the estimated duration of 
time required for participation in the study, as well as alternative interventions or courses of 
treatment, if applicable.   
 
The subject and parent/legal guardian must receive an explanation as to whether any 
compensation and any medical treatments are available if injury occurs and, if so, what they are, 
where further information may be obtained, and who to contact in the event of a study-related 
injury.  Subject and parent/legal guardian must be told who to contact for answers to any 
questions related to the study.  The extent of the confidentiality of subject records must be 
defined and the subject must be informed that applicable data protection legislation applies. 
 
The subject and parent/legal guardian must be informed that the monitor(s), auditor(s), IRB/IEC 
members, and the applicable regulatory authorities will be granted direct access to the subject’s 
original study medical records for verification of protocol-specified procedures and/or data, 
without violating the confidentiality of the subject to the extent permitted by the applicable laws 
and regulations.  The subject and parent/legal guardian must be informed that his/her signature 
on the informed assent or consent form indicates that he/she has decided to participate in the 
study, having read and discussed the information presented.  
 


 
 
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Modifications made by the investigator to an informed assent or consent form template provided 
to the investigator by Aeras or its designee will be reviewed and approved by Aeras or its 
designee prior to being submitted to the IRB/IEC.   
 
The original, signed informed assent and consent forms for each subject and parent/legal 
guardian will be maintained by the investigator as part of the subject’s study records.  A copy of 
the signed informed assent and consent forms will be provided to each subject and parent/legal 
guardian. 
 
10 
STUDY COMPLETION 
At the discretion of Aeras, all materials and supplies provided to the investigator will be returned 
or disposed of in compliance with local regulatory requirements upon authorization from Aeras, 
upon study completion.  The investigator or designated clinical site staff will notify the IRB/IEC 
when the study has been completed. 
 
11 
PUBLICATIONS 
The final study report will be made available to the principal investigator for purposes of 
publications.  The principal investigator and study staff must send all manuscripts, abstracts, and 
presentations using data from this study to Aeras for review prior to their submission.  Aeras 
reserves the right to delete any part or parts of such materials deemed to be confidential or 
proprietary. 
 
12 
CHANGES IN THE PROTOCOL 
The protocol may not be modified without written approval from Aeras.  All changes to the 
protocol must be submitted to the IRB/IEC and must be approved by the IRB/IEC prior to their 
implementation.   
 
12.1 
Changes from Version 1.0 to Version 2.0 (Amendment 1) 
Section Number 
(Title) 
Change/Rationale 
Section 3.1 
(Schedule of 
Subject 
Evaluations) 
Tables 3-1 and 3-2 were revised as follows: 
• For the Correlates Cohort, collection of solicited AEs (through Study Day 7) and 
unsolicited AEs (through Study Day 28) were added to the applicable visits. This 
change was made to collect more safety information since this study represents the 
first use of AERAS-404 in adolescents. 
• Blood collection (0.5 mL) for absolute blood count was added as follows: for the 
Correlates Cohort at Study Days 0 and 3 (AERAS-404/placebo) and Study Days 0 
and 7 (BCG); and for the Safety & Immunogenicity Cohort and Correlates Cohort at 
Months 6, 12, 18, and 24. These changes were made in order to have samples 
collected for absolute blood counts at the same time points as for RNA analysis, to 
deconvolute the RNA results at all time points. 
• Collection of serum for correlates of risk/protection was added at Study Days 0 and 
70, and Months 6, 12, 18, and 24. This was done at the request of MCC to obtain 
antigen specific antibodies. 


 
 
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• A footnote was added to indicate that all blood volumes are approximate. 
• A typographical error (Table 3-2 only) that indicated that the Day 3 visit is for 
subjects in the Safety & Immunogenicity Cohort only was corrected. In fact, all 
subjects who receive AERAS-404 or placebo will have a visit at Day 3. 
Section 3.2.4 
(Exclusion 
Criteria) 
Exclusion criterion #2 was revised to refer to axillary temperature instead of oral 
temperature, to maintain consistency with the protocol toxicity table in Appendix C. 
 
Tracked changes: 
2.  OralAxillary temperature 37.5C on Study Day 0 
Section 3.4.1 
(Unblinding for 
Clinical 
Emergencies) 
Change/rationale:  The description for requesting unblinding of a subject’s treatment 
assignment in the event of urgent clinical requirement was corrected to reflect the 
process used with the IWRS. 
 
Tracked changes: 
‘If there is an urgent clinical requirement to know a subject’s treatment assignment, the 
principal investigator will request the urgent unblinding of a subject’s treatment by 
following the “unblinding by site form call flow” processcompleting the Subject 
Unblinding by Site form in the IWRS.’ 
Section 3.6.2 
(Immunology 
Laboratory 
Evaluations) 
Table 3-3 was revised as follows: 
• To reflect the changes made to the schedule of evaluation Tables 3-1 and 3-2 (see 
Changes for Section 3.1, above) 
• To indicate that the volume of blood to be collected for ‘PBMC, plasma, and serum’ 
at Study Days 0 and 70 will be ‘up to 16mL’ (previously noted as ‘16mL’). 
• To indicate in the ‘PBMC, plasma, and serum’ row that 2mL (previously 1mL) of 
plasma will be collected at each time point, and that 1 clotted tube will be used (in 
addition to the CPT tubes previously noted). 
• To correct the volume of blood to be collected for ‘PBMC, plasma, and serum’ at 
Months 6, 12, 18, and 24 to read ‘up to 40mL’ (previously was noted as ‘16mL’). 
• To delete the row for ‘Whole blood in PAXgene’ under ‘Early innate signature 
(Safety & Immunogenicity Cohort only, n=90).’ This row was previously included in 
error, and represented a duplicate of information included under ‘Correlates of 
Protection (Safety & Immunogenicity and  Correlates cohorts , n=990).’  
Section 3.6.3.3 
(Adverse Events) 
Table 3-4 was revised to reflect that unsolicited and solicited adverse events will now be 
collected for the Correlates Cohort, and that the diary card for solicited AEs will only be 
used for the Safety and Immunogenicity Cohort. 
Section 4.1 
(Supplies) 
This section was revised to indicate that the normal saline placebo will be sourced as 
either ampoules or vials (previously only vials were indicated). 
Section 4.3 
(Receipt and 
Storage) 
This section was revised to indicate that AERAS-404 must be stored at ‘less than or 
equal to -15C’ (previously ‘-15C’ was indicated). 
Section 5.12 
(Reporting of 
Serious Adverse 
Events) 
This section was revised to reflect the method of reporting serious adverse events, and 
for clarity. 
 
Tracked changes: 
All seriousSerious adverse events, which include SUSARs, are reported to the sponsor 
and to the World Wide Safety Center (administered by PPD, Inc.) for the entire study 
period (see protocol appendices).  SUSARs are reported even after the trial is over, if the 
sponsor, local medical monitor or principal investigator becomes aware of them.  The 
site will be provided with specific reporting procedures including the SAE eCRFAdverse 
Event CRF and any supplemental reporting forms to be used.  Serious adverse events 
will be reported on the SAE eCRFAdverse Event CRF using a recognized medical term 


 
 
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or diagnosis that accurately reflects the event.   
 
Serious adverse events will be assessed for severity, causal relationship to the study 
vaccine, and expectedness by the investigator and the local medical monitor according to 
their roles (as described in Sections 5.1.1 and 5.1.3) for severity, causal relationship to 
the study vaccine, and expectedness.  The onset and resolution dates of the event and 
medical carethe action taken in response to the event will be documented.  If the event 
has not resolved by the final study visit, it will be documented as “ongoing” on the 
eCRF, however, follow-up of the SAE must continue until resolved or the condition has 
stabilized.  Information recorded on the eCRF must be substantiated in the source 
documents. 
 
The SAE eCRFReport form completed for that event must be completedscanned and 
emailed, or faxed by the principal investigator or his/her designee, within 24 hours (one 
businesscalendar day) of the clinical site becoming aware of the event, to the local 
medical monitor and to the World-Wide Safety Center at PPD.  The SAE eCRF should 
be completed with all information known at the time;. In case the eCRF cannot be 
completed, the Supplemental SAE Report (paper form) should be completed by the 
principal investigator or his/her designee, and both forms scanned and emailed, or faxed 
to(even if all information concerning the World Wide Safety Center and to the local 
medical monitorevent is not yet known) within the first 24 hours of awareness of the 
event.    
 
.... 
 
Aeras has authorized the PPD World Wide Safety Center to execute its responsibilities 
for expedited safety report submission to the appropriate regulatory authorities within 
specific time periods of being notified of the event (within 7 or 15 calendar days 
depending the character of the SUSAR); therefore, it is important that the investigator 
submit additional information requested as soon as it becomes available.   
 
Aeras will notify the DMC and the manufacturer of AERAS-404 of all SUSARs within 3 
working days of becoming aware of an event and will provide all follow-up information 
in a timely manner. 
Section 7.3 
(Prevention of 
Infection (QFT-
GIT Conversion) 
Analyses) 
A typographical error regarding reversion of QFT-GIT conversion was corrected. 
 
Tracked changes: 
The methods used for the analysis of this endpoint will be the same as that for the 
primary Mtb infection endpoint, but the number of endpoints will be smaller as subjects 
who experience primary QFT-GIT conversion but then have a reversion to 
negativepositive at one or both of the repeat tests will not be counted. 
Appendix A 
(Detailed 
Description of 
Study Visits) 
The visit descriptions were revised as follows: 
• To reflect the changes made to the schedule of evaluation Tables 3-1 and 3-2 (see 
Changes for Section 3.1, above). 
• To change oral temperature to axillary temperature, to maintain consistency with the 
protocol toxicity table in Appendix C. 
• To remove urine collection from the Study Day 3 visit for subjects receiving AERAS-
404 or placebo in the Safety and Immunogenicity Cohort, for consistency with Table 
3-2 
• To correct the window for the Month 6, 12, 18, and 24 visits for subjects receiving 
AERAS-404 or placebo to read ‘±14 days from Day 0’ (previously read ‘±14 days 
from Day 56’). 


 
 
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Appendix C 
(Toxicity Table 
(Modified from 
Division of AIDS 
Table for Grading 
the Severity of 
Adult and Pediatric 
Adverse Events)) 
The following changes were made to the definitions in the toxicity table: 
• 
One of the definitions for Grade 3 hemoglobin (any decrease) was corrected to read 
‘≥2.79 mmol/L’ (previously read ‘>2.79 mmol/L’). 
• 
The definitions for Grade 1 and Grade 2 for hematuria (microscopic) in the toxicity 
table were changed to reflect the definitions used at the site. 
Tracked changes: 
Grade 1: 65 – 10 cells/mm3RBC/HPF 
Grade 2: > 10 cells/mm3RBC/HPF 
Appendix D 
(Adverse Events of 
Special Interest) 
Autoimmune thrombotic/thromboembolic conditions and Tolosa-Hunt syndrome were 
added to the list of adverse events of special interest. This was done at the request of the 
DMC to fully assess the safety profile of the vaccine. 
Throughout the 
protocol 
Minor editorial changes were made for clarity. 
 
12.2 
Changes from Version 2.0 to Version 3.0 (Amendment 2) 
Section Number 
(Title) 
Change/Rationale 
Study Abstract 
The study abstract was revised the reflect the changes below. 
Section 2.2 
(Design and 
Endpoints) 
The first 2 paragraphs of this section were revised to reflect that additional sites to 
SATVI may be used, and that subjects will be recruited directly from the community (in 
addition to high schools). 
Section 3.1 
(Schedule of 
Subject 
Evaluations) 
Tables 3-1 and 3-2 were revised as follows: 
• Written informed consent and assent were removed from the screening procedures 
(text was added above the tables to indicate that written informed consent and assent 
must be obtained before any screening procedures are performed). 
• Whole blood collection for evaluation of novel IGRA was added at screening. 
• A window of ±14 days was added for each of the post QFT-GIT(-) to QFT-GIT(+) 
conversion visits. 
Section 3.2.1 
(Recruitment and 
Informed Consent) 
The first paragraph of this section was revised to reflect that subjects will be recruited 
directly from the community (in addition to high schools), and to add community 
organizations to the list of groups used to recruit subjects. 
Section 3.2.3 
(Inclusion Criteria) 
Inclusion criterion #6 was deleted (‘Has body mass index [BMI] for age and sex between 
the 5th and 95th centiles by Centers for Disease Control nomogram’). This criterion has 
resulted in the exclusion of approximately 15% of the first 200 participants screened who 
are otherwise healthy, with exclusions occurring both above and below the 5th/95th 
centiles. SATVI data indicate that BMI in this population has a bimodal distribution, 
with peaks at the high and low ends of the curve. Thus the use of a BMI nomogram does 
not appear to be appropriate for this population and excludes otherwise eligible 
participants without increasing safety. 
Section 3.2.4 
(Exclusion 
Criteria) 
A new exclusion criterion was added to prevent the possibility of immunological 
interference by use of licensed vaccines in close temporal proximity to study vaccine 
administration: 
‘14 Planned administration/administration of a licensed vaccine in the period starting 28 
days before and ending 28 days after each dose of study vaccine’. 
Section 3.3 
(Study 
Randomization) 
This section was revised to reflect that additional sites to SATVI may be used. 
 
Tracked Changes: 
....The day of randomization for each subject will be Study Day 0. Randomization will be 
in blocks for each school or community, as the epidemiology of TB transmission may 
vary from one school or community to another. Randomization will also be blocked such 


 
 
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that 30 subjects from each vaccine group will be enrolled in the Safety & 
Immunogenicity Cohort. Enrollment for the Safety & Immunogenicity Cohort will be 
from a subset of schools or communities selected to be representative of the study 
population as a whole in terms of socio-economic status and TB risk. Subjects who 
discontinue participation will not be replaced. 
Section 3.6.1 
(Evaluation of Mtb 
Infection) 
The text regarding performance of QFT-GIT assays was clarified to indicate that the 
assay will be performed according to site SOPs.  
Section 3.6.2 
(Immunology 
Laboratory 
Evaluations) 
Table 3-3 was revised to indicate that the secondary immunogenicity ICS assay and 
absolute blood cell subset counting by flow cytometry will be conducted at the 
applicable site laboratory (previously the SATVI laboratory was listed as conducting 
both of these assays). 
Section 3.6.3.3 
(Adverse Events) 
Table 3-4 was revised to include the following clarifications regarding the collection 
periods for adverse events: 
• 
The 28-day post-vaccination collection period for unsolicited AEs and 7-day post-
vaccination period for solicited AEs refer to time periods after each vaccination 
• 
Solicited injection site reaction AEs will be collected for 84 days post-vaccination in 
the BCG group and 28 days post each vaccination in the AERAS-404 and placebo 
groups 
• 
For both unsolicited AEs and solicited injection site AEs, events will be collected 
through the end of the study visit window for each adverse event collection period 
(i.e., 28 days after each vaccination + 5-day window = 33 days) 
Section 5.1.1 
(Principal 
Investigator) 
The following text was added to define designee for the principal investigator:  
‘Where specified, the responsibilities of the principal investigator may be delegated to a 
medically qualified team member (designee).’  
Subsequent text in this section was revised to indicate that a designee for the principal 
investigator can determine the severity and causality with respect to study vaccine of 
adverse events. 
Section 5.3 
(Definition of 
Adverse Event) 
The text in this section was revised to indicate that a designee for the principal 
investigator may estimate the resolution date of an adverse event if the resolution date is  
uncertain. 
Section 5.7 
(Solicited Adverse 
Events and 
Injection Site 
Reactions) 
The following text was added to this section to describe the circumstances under which 
injection site reactions should be photographed:  
‘Visible injection-site abnormalities other than expected site reactions should be 
photographed. The types of events to be photographed should be findings that are grossly 
abnormal or beyond the expected injection site reactions. These types of reactions may 
be reactions that appear rapidly or are larger or more severe than usual. The purpose of 
the photograph is for information sharing purposes with the sponsor. The types of events 
that should be photographed include, but are not limited to, allergic type reactions, such 
as prompt injection site redness, angioedema, and anaphylactic reaction.’ 
Appendix A 
(Detailed 
Description of 
Study Visits) 
The visit descriptions were revised for consistency with the changes described above, 
and to add timing for the injection site examinations on the day of each injection (30±5 
minutes post-immunization). 
Appendix C 
(Toxicity Table 
(Modified from 
Division of AIDS 
Table for Grading 
the Severity of 
Adult and Pediatric 
Adverse Events)) 
The definitions for pediatric (≤17 years) hypertension were revised to add values 
required for systolic and diastolic blood pressure to meet the criteria for a moderate or 
severe AE. 


 
 
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12.3 
Changes from Version 3.0 to Version 4.0 (Amendment 3) 
Section Number 
(Title) 
Change/Rationale 
Section 2.2 
(Design and 
Endpoints) 
A footnote was added to Table 2-1 to indicate that subject numbers are approximate due 
to block randomization. 
Section 3.1 
(Schedule of Subject 
Evaluations) 
Tables 3-1 and 3-2 were revised as follows: 
• Study Days were defined for the Month 6, 12, 18, and 24 visits, and the numbers of 
days post conversion were added for the 3-month and 6-month post-QFT-GIT-
conversion visits, for clarification of timing. 
• HIV-1 testing was revised to indicate that this includes HIV counselling. 
• Collection of injection site reaction adverse events was revised to include unsolicited 
events (previously was only solicited events). 
• PBMC, plasma, and serum at Months 18 and 24 for correlates of protection were 
removed. This was done as the most useful correlates at the later time points are 
likely to be those determined from the absolute blood count and RNA.  
• Blood for absolute blood count and RNA was added at the visits after QFT 
conversion, to explore correlates of risk for TB disease.  
Section 3.2.4 
(Exclusion Criteria) 
The following changes were made to the exclusion criteria: 
• 
Exclusion criterion #3 was revised to clarify the timing of blood collection for 
qualifying laboratory tests, and to add more specific criteria for each laboratory 
parameter that would result in exclusion of a subject from the study. 
 
Tracked Changes: 
Clinically significant (and no more than Grade 1 on the Toxicity Scale) aAbnormal 
laboratory values from the most recent blood collected within 21 days prior to 
randomization as follows: 
• 
Laboratory evidence of hematologic disease (white blood cell count <3000/mm3 
or >11,500/mm3; hemoglobin <0.9 times the lower limit of normal of the testing 
laboratory, by age and gender; absolute neutrophil count <1300/mm3; absolute 
lymphocyte count <1000/mm3)hemoglobin, hematocrit, absolute neutrophil 
count, or absolute lymphocyte count below lower limit of normal (LLN)  
• 
white blood cell count above upper limit of normal (ULN) or below LLN (i.e., 
must be within normal limits)  
• 
ALT, AST, alkaline phosphatase, total bilirubin, creatinine, blood urea nitrogen 
(BUN) >1.25 times the upper limit of normal of the testing laboratoryabove 
ULN  
 
• 
Exclusion criterion #4 was revised to clarify exclusion based on urinalysis results. 
 
Tracked Changes: 
Evidence of clinically significant (and no more Greater than Grade 1 on the Toxicity 
Scale) systemic or local disease on urinalysis result (with the exception of hematuria 
in a menstruating female), or urinalysis abnormality judged clinically significant by 
the investigator 
Section 3.6.2 
(Immunology 
Evaluations) 
Table 3-3 was revised to reflect the changes made to the schedule of evaluation Tables 3-
1 and 3-2 (see Changes for Section 3.1, above). In addition, absolute blood count and 
RNA will be collected at visits after QFT conversion, to explore correlates of risk.  
Section 3.6.3.3 
Adverse Events 
Table 3-4 was revised to indicate that collection of injection site reaction adverse events 
includes unsolicited and solicited adverse events (previously was only solicited adverse 
events). 


 
 
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Section 3.6.3.4 
Concomitant 
Medications 
This section was revised to clarify the collection periods for concomitant medications, 
and to indicate that TB medications will be collected throughout the study. 
Section 5.1.4 
(Data Monitoring 
Committee) 
This section was revised as follows in relation to DMC review of unblinded QFT-GIT 
conversion endpoints when sufficient endpoints have accumulated for preliminary 
analysis: 
• 
To clarify that when the DMC advises the Sponsor after this review, both positive 
and negative impact on infection will be taken into account. 
• 
To indicate that as an alternative to electing to extend follow-up based on this 
review, the Sponsor may elect to terminate the study. 
Section 7.4.2.1 
(RNA 
Transcriptomics and 
Whole Blood 
Absolute Blood Cell 
Counts) 
The following text was added to this section: 
‘Samples collected after QFT conversion will be used to explore a correlate of risk for 
developing TB disease which has been identified by SATVI, or to explore other 
correlates as informed by recent data.’ 
Section 7.4.2.3 
Correlates of 
Risk/Correlates of 
Protection 
This section was revised to reflect the changes made to the schedule of evaluation Tables 
3-1 and 3-2 (see Changes for Section 3.1, above). 
Appendix A 
(Detailed 
Description of Study 
Visits) 
The visit descriptions were revised for consistency with the changes described above and 
for consistency with Schedule of Subject Evaluation Tables 3-1 and 3-2. 
 
12.4 
Changes from Version 4.0 to Version 5.0 (Amendment 4) 
Section Number 
(Title) 
Change/Rationale 
Section 2.1 
(Objectives) 
A new exploratory objective was added to evaluate early QFT converters (i.e., those that 
converted at Month 6 or 12) for sustained QFT conversions or late reversions so as to 
inform the design of subsequent POI studies. 
Added: 
To explore trends in QFT-GIT prolonged/sustained conversions and late reversions (i.e., 
more than 6 months post initial conversion) in early QFT-GIT converters (i.e., among 
those who converted at Month 6 or Month 12 of follow-up) 
Section 2.2 
(Design and 
Endpoints), 
Section 3.1 
(Schedule of Subject 
Evaluations), 
Section 3.6.2 
(Immunology 
Laboratory 
Evaluations), 
and Appendix A 
(Detailed 
Description of 
Subject Visits) 
These sections were updated to include the optional, additional visit for participants who 
converted to QFT-GIT+ at Month 6 or Month 12. 
 
 


 
 
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Section 3.2.1 
(Recruitment and 
Informed Consent) 
A requirement was added to obtain consent/assent for early QFT converters to participate 
in the optional, additional follow-up visit described above. 
Added: 
Subjects who converted to a positive QFT-GIT at Month 6 or 12 will be approached for 
an additional QFT-GIT test, blood collection for absolute blood count and RNA, and 
evaluation for signs/symptoms of TB to be conducted at least 24 months after their initial 
vaccination. Informed consent will be obtained by the use of a written supplemental 
consent form approved by the IRB or IEC, and signed and dated by subject or by the 
subject’s parent/legal guardian if subject is less than 18 years of age.  Similarly, 
informed assent will be obtained by the use of a written assent form approved by the IRB 
or IEC, and signed and dated by the subject if the subject is less than 18 years of age. 


 
 
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13 
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adjuvanted with IC31 promotes strong and long-lived Mycobacterium tuberculosis specific T 
cell responses in naïve human volunteers. Vaccine. Apr 30;28(20):3571-81. 
 


 
 
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Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
van Dissel JT, Soonawala D, Joosten SA, Prins C, Arend SM, et al.  (2011) Ag85B-ESAT-6 
adjuvanted with IC31® promotes strong and long-lived Mycobacterium tuberculosis specific T 
cell responses in volunteers with previous BCG vaccination or tuberculosis infection. Vaccine. 
Mar 3;29(11):2100-9.  
 
van Zyl-Smit RN, Pai M, Peprah K, Meldau R, Kieck J, et al. (2009) Within-subject variability 
and boosting of T-cell interferon-gamma responses after tuberculin skin testing. Am J Respir Crit 
Care Med 180: 49-58. 
 
WHO, Global Tuberculosis Report 2012. http://www.who.int/tb/publications/global_report/en/ 
 
Wilson ME, Fineberg HV, Colditz GA. (1995) Geographic latitude and the efficacy of bacillus 
Calmette-Guerin vaccine. Clin Infect Dis 20:982–991. 
 
Zwerling A, Benedetti A, Cojocariu M, McIntosh F, Pietrangelo F, et al. (2013) Repeat IGRA 
testing in Canadian health workers: conversions or unexplained variability? PLoS One 8: 
e54748. 


 
 
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APPENDIX A 
Detailed Description of Study Visits 
 
Screening Visit(s)  (All Subjects) 
Written informed consent and assent must be obtained before any screening procedures are 
performed. 
 
1. Verify study entry eligibility criteria are met 
2. Medical history 
3. Physical examination  
4. Urine collection for urinalysis and (for females) HCG 
5. Vital Signs 
6. TB Symptom Screen 
7. Blood collection for  
• 
QuantiFERON®-TB Gold In-Tube 
• 
HIV-1 (with pre- and post-test counselling) 
• 
Serum chemistry (AST, ALT, alkaline phosphatase, total bilirubin, creatinine, BUN) 
• 
Hematology (hemoglobin, hematocrit, white cell count with differential and platelet count) 
• 
Whole blood for novel IGRA 
 
 
Study Day 0 (All Subjects) 
 
Pre- injection: 
1. Urine collection (females) for HCG 
2. Obtain pre-immunization vital signs (blood pressure, pulse, axillary temperature)  
3. Interval medical history and, if indicated by history, focused physical examination. 
4. Verify study entry eligibility criteria are met 
5. Randomize 
6. Blood collection for 
• 
PMBC and plasma for correlates of risk protection 
• 
Whole blood for RNA extraction/absolute blood count 
• 
Whole blood for novel IGRA 
• 
PBMC for evaluation of NTM exposure 
 
Safety & Immunogenicity Cohort Only: 
• 
Whole blood assay 
• 
PMBC for ICS 
• 
Serum for multiplex ELISA 
 
Injection: 
7. Administer either 1) BCG by intradermal injection or 2) AERAS-404 or placebo by intramuscular injection on 
the left upper arm (deltoid region).  Record date and time of injection and which arm was injected. 
 
Post-injection: 
8. Monitor participant for solicited and unsolicited adverse events for at least 30 minutes after injection; record 
concomitant medications 
9. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs for at least 30 
minutes after injection; record concomitant medications 
10. Obtain 305-minute post-immunization vital signs  


 
 
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11. Complete 305-minute post-immunization Study Day 0 study injection site examination (with photograph if 
there is a visible injection-site abnormality; see Section 5.7 for further details) 
 
Safety & Immunogenicity Cohort Only: 
12. Distribute diary cards 
 
 
Post Study Day 0 Visits (Subjects receiving BCG) 
 
Study Day 7 (Subjects receiving BCG) 
Allowable window for clinic visit is 7±1 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for solicited and unsolicited adverse events; record concomitant medications 
3. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
4. Monitor participant for solicited and unsolicited injection site reaction adverse events; record concomitant 
medications 
5. Complete Study Day 0 injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
6. Whole blood collection for RNA extraction/absolute blood count 
 
Safety & Immunogenicity Cohort only 
7. Urine collection for urinalysis 
8. Review diary cards. 
9. Blood collection for 
• 
Serum chemistry (AST, ALT, alkaline phosphatase, total bilirubin, creatinine, BUN) 
• 
Hematology (hemoglobin, hematocrit, white cell count with differential and platelet count) 
• 
Serum for multiplex ELISA 
 
Study Day 28 (Subjects receiving BCG) 
Allowable window for clinic visit is 28±5 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for unsolicited adverse events; record concomitant medications 
3. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
4. Monitor participant for solicited and unsolicited injection site reaction adverse events; record concomitant 
medications 
5. Complete Study Day 0 study injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
6. TB Symptom Screen 
 
Study Day 70 (Subjects receiving BCG) 
Allowable window for clinic visit is 70±5 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
3. Monitor participant for solicited and unsolicited injection site reaction adverse events; record concomitant 
medications 
4. Complete Study Day 0 study injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
5. TB Symptom Screen 
6. Blood collection for  


 
 
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• 
PBMC and plasma for correlates of risk/protection  
 
Safety & Immunogenicity Cohort only 
7. Blood collection for  
• 
Whole blood assay 
• 
PBMC for ICS 
 
Study Day 84 (Subjects receiving BCG) 
Allowable window for clinic visit is 84±7 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
3. Monitor participant for solicited and unsolicited injection site reaction adverse events; record concomitant 
medications 
4. Complete Study Day 0 study injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
5. TB Symptom Screen 
6. Blood collection 
• 
QuantiFERON®-TB Gold In-Tube 
• 
Whole blood for novel IGRA 
 
 
Study Month 6, 12, 18 and 24 (Study Days 168, 336, 504, and 672) (Subjects receiving BCG; 
to be completed by subjects who did not convert from QFT-GIT(-) to QFT-GIT(+) at the 
previous visit) 
Allowable window for clinic visit is 6, 12, 18, or 24 months (168, 336, 504, 672 days) ±14 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
3. TB Symptom Screen 
4. Blood collection 
• 
QuantiFERON®-TB Gold In-Tube 
• 
Whole blood for novel IGRA 
• 
[Study Months 6 and 12 only]PBMC and plasma for correlates of risk protection 
• 
Whole blood for RNA extraction/absolute blood count 
 
Study Month 6 (Study Day 168) (Subjects receiving BCG; to be completed by subjects who 
converted from QFT-GIT(-) to QFT-GIT(+) at Study Day 84) 
Allowable window for clinic visit is 6 months (168 days) ±14 days from Day 0 
1. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
2. TB symptom screen 
 
84 and 168 Days after conversion from QFT-GIT(-) to QFT-GIT(+) at Months 6-24 
(Subjects receiving BCG; to be completed by subjects who converted from QFT-GIT(-) to 
QFT-GIT(+) at Month 6-24 visits) 
Allowable window for clinic visit is 84 or 168 days ±14 days after conversion 
1. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
2. TB symptom screen 
3. Blood collection 
• 
QuantiFERON®-TB Gold In-Tube  


 
 
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• 
Whole blood collection for RNA extraction/absolute blood count 
 
Final visit for subjects who converted from QFT-GIT(-) to QFT-GIT(+) at Month 6 or 12 
(Subjects receiving BCG) 
Written informed consent and assent must be obtained for additional follow-up before any 
extended follow-up procedures are performed. 
Allowable window for clinic visit is at least 24 months after initial vaccination but before the end of the study  
1. TB symptom screen 
2. Record TB treatment, if applicable 
3. Blood collection 
• 
QuantiFERON®-TB Gold In-Tube 
• 
Whole blood for RNA extraction/absolute blood count 
 
Post Study Day 0 Visits (Subjects receiving AERAS-404 or placebo) 
 
Study Day 3 (Subjects receiving AERAS-404 or placebo) 
Allowable window for clinic visit is 3±1 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for solicited and unsolicited adverse events; record concomitant medications 
3. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
4. Monitor participant for solicited injection site reaction adverse events at Study Day 0 injection site; record 
concomitant medications 
5. Complete Study Day 0 injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
6. Whole blood collection for RNA extraction/absolute blood count 
 
Safety & Immunogenicity Cohort only 
7. Blood collection for 
• 
Serum for multiplex ELISA 
 
Study Day 7 (Subjects receiving AERAS-404 or placebo) 
Allowable window for clinic visit is 7±1 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for solicited and unsolicited adverse events; record concomitant medications 
3. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
4. Monitor participant for solicited injection site reaction adverse events at Study Day 0 injection site; record 
concomitant medications 
5. Complete Study Day 0 injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
 
Safety & Immunogenicity Cohort only 
6. Urine Collection for urinalysis 
7. Review diary cards. 
8. Blood collection for 
• 
Serum chemistry (AST, ALT, alkaline phosphatase, total bilirubin, creatinine, BUN) 
• 
Hematology (hemoglobin, hematocrit, white cell count with differential and platelet count) 
 
Study Day 28 (Subjects receiving AERAS-404 or placebo) 
Allowable window for clinic visit is 28±5 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 


 
 
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2. Monitor participant for unsolicited adverse events; record concomitant medications 
3. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
4. Monitor participant for solicited injection site reaction adverse events at Study Day 0 injection site; record 
concomitant medications 
5. Complete Study Day 0 injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
6. TB Symptom Screen 
 
Study Day 56 (Subjects receiving AERAS-404 or placebo) 
Allowable window for clinic visit is 56-2/56+14 days from day 0 
 
Pre- injection: 
1. Urine collection (females) for HCG 
2. Obtain pre-immunization vital signs (blood pressure, pulse, axillary temperature)  
3. Interval medical history and, if indicated by history, focused physical examination. 
4. Verify study entry eligibility criteria are met 
 
Injection: 
5. Administer AERAS-404 or placebo by intramuscular injection on the right upper arm (deltoid region).  Record 
date and time of injection and which arm was injected. 
 
Post-injection: 
6. Monitor participant for solicited and unsolicited adverse events for at least 30 minutes after injection; record 
concomitant medications 
7. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs for at least 30 
minutes after injection; record concomitant medications 
8. Monitor participant for solicited injection site reaction adverse events at Study Day 56 injection site; record 
concomitant medications 
9. Obtain 305-minute post-immunization vital signs 
10. Complete 305-minute post-immunization Study Day 56 study injection site examination (with photograph if 
there is a visible injection-site abnormality; see Section 5.7 for further details) 
11. TB symptom screen 
 
Safety & Immunogenicity Cohort only 
12. Distribute diary cards 
 
Study Day 63 (Subjects receiving AERAS-404 or placebo) 
Allowable window for clinic visit is 7±1 days from Day 56 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for solicited and unsolicited adverse events; record concomitant medications 
3. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
4. Monitor participant for solicited injection site reaction adverse events at Study Day 56 injection site; record 
concomitant medications 
5. Complete Study Day 56 injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
 
Safety & Immunogenicity Cohort only 
6. Urine Collection for urinalysis 
7. Review diary cards. 
8. Blood collection for 


 
 
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• 
Serum chemistry (AST, ALT, alkaline phosphatase, total bilirubin, creatinine, BUN) 
• 
Hematology (hemoglobin, hematocrit, white cell count with differential and platelet count) 
 
Study Day 70 (Subjects receiving AERAS-404 or placebo) 
Allowable window for clinic visit is 14±3 days from Day 56 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for unsolicited adverse events; record concomitant medications 
3. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
4. Monitor participant for solicited injection site reaction adverse events at Study Day 56 injection site; record 
concomitant medications 
5. Complete Study Day 56 injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
6. Blood collection for 
• 
PBMC and plasma for correlates of risk/protection  
 
Safety & Immunogenicity Cohort only 
7. Blood collection for 
• 
Whole blood assay 
• 
PMBC for ICS 
 
Study Day 84 (Subjects receiving AERAS-404 or placebo) 
Allowable window for clinic visit is 28±5 days from Day 56 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for unsolicited adverse events; record concomitant medications 
3. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
4. Monitor participant for solicited injection site reaction adverse events at Study Day 56 injection site; record 
concomitant medications 
5. Complete Study Day 56 injection site examination (with photograph if there is a visible injection-site 
abnormality; see Section 5.7 for further details) 
6. TB symptom screen 
7. Blood collection for  
• 
QuantiFERON®-TB Gold In-Tube  
• 
Whole blood for evaluation of IGRA 
 
Study Month 6, 12, 18 and 24 (Study Days 168, 336, 504, and 672) (Subjects receiving 
AERAS-404 or placebo; to be completed by subjects who did not convert from QFT-GIT(-) 
to QFT-GIT(+) at the previous visit) 
Allowable window for clinic visit is 6, 12, 18, or 24 months (168, 336, 504, 672 days) ±14 days from Day 0 
1. Interval medical history and, if indicated by history, focused physical examination. 
2. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; and record 
concomitant medications 
3. TB Symptom Screen 
4. Blood collection for  
• 
QuantiFERON®-TB Gold In-Tube  
• 
Whole blood for evaluation of IGRA 
• 
[Study Months 6 and 12 only]PBMC and plasma for correlates of risk protection 
• 
Whole blood for RNA extraction/absolute blood count 
 


 
 
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Study Month 8 (Study Day 224) (Subjects receiving AERAS-404 or placebo; to be 
completed by subjects who converted from QFT-GIT(-) to QFT-GIT(+) at Study Day 84) 
Allowable window for clinic visit is 6 months (168 days) ±14 days from Day 56 
1. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
2. TB symptom screen 
 
84 and 168 Days after conversion from QFT-GIT(-) to QFT-GIT(+) at Months 6-24 
(Subjects receiving AERAS-404 or placebo; to be completed by subjects who converted 
from QFT-GIT(-) to QFT-GIT(+) at Month 6-24 visits) 
Allowable window for clinic visit is 84 or 168 days ±14 days after conversion 
1. Monitor participant for serious adverse events, adverse events of special interest, and SUSARs; record 
concomitant medications 
2. TB symptom screen 
3. Blood collection 
• 
QuantiFERON®-TB Gold In-Tube  
• 
Whole blood collection for RNA extraction/absolute blood count 
 
Final visit for subjects who converted from QFT-GIT(-) to QFT-GIT(+) at Month 6 or 12 
(Subjects receiving AERAS-404 or placebo)  
Written informed consent and assent must be obtained for additional follow-up before any 
extended follow-up procedures are performed. 
Allowable window for clinic visit is at least 24 months after initial vaccination but before the end of the study  
1. TB symptom screen 
2. Record TB treatment, if applicable 
3. Blood collection  
• 
QuantiFERON®-TB Gold In-Tube 
• 
Whole blood for RNA extraction/absolute blood count  
 


 
 
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SAE and SUSAR Reporting Schemes 
 
 
 
 
 
 
 
 


 
 
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APPENDIX B 
Toxicity Table (Modified from Division of AIDS Table for 
Grading the Severity of Adult and Pediatric Adverse Events) 
 
PARAMETER 
GRADE 1 
MILD 
GRADE 2 
MODERATE 
GRADE 3 
SEVERE 
GRADE 4 
POTENTIALLY 
LIFE-THREATENING 
ESTIMATING SEVERITY GRADE 
Clinical adverse event 
NOT identified 
elsewhere in this AE 
Grading Table  
Symptoms causing 
no or minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
greater than minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
inability to perform 
usual social & 
functional activities 
Symptoms causing 
inability to perform 
basic self-care functions 
OR Medical or 
operative intervention 
indicated to prevent 
permanent impairment, 
persistent disability, or 
death  
SYSTEMIC 
Acute systemic 
allergic reaction  
Localized urticaria 
(wheals) with no 
medical intervention 
indicated 
Localized urticaria 
with medical 
intervention indicated 
OR Mild angioedema 
with no medical 
intervention indicated 
Generalized urticaria 
OR Angioedema with 
medical intervention 
indicated OR 
Symptomatic mild 
bronchospasm 
Acute anaphylaxis OR 
Life-threatening 
bronchospasm OR 
laryngeal edema 
Chills 
Symptoms causing 
no or minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
greater than minimal 
interference with 
usual social & 
functional activities  
Symptoms causing 
inability to perform 
usual social & 
functional activities 
NA 
Fatigue 
Malaise 
Symptoms causing 
no or minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
greater than minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
inability to perform 
usual social & 
functional activities 
Incapacitating fatigue/ 
malaise symptoms 
causing inability to 
perform basic self-care 
functions 
Fever (axillary) 
38.0 – 38.4C 
100.4 – 101.1F 
38.5 - 40C 
101.2 - 104F 
>40C 
>104F 
N/A 
Pain (indicate body 
site) 
DO NOT use for pain 
due to injection (See 
Injection Site 
Reactions: Injection 
site pain) 
See also Headache, 
Arthralgia, and 
Myalgia 
Pain causing no or 
minimal interference 
with usual social & 
functional activities 
Pain causing greater 
than minimal 
interference with 
usual social & 
functional activities 
Pain causing inability to 
perform usual social & 
functional activities 
Disabling pain causing 
inability to perform 
basic self-care functions 
OR Hospitalization 
(other than emergency 
room visit) indicated 


 
 
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PARAMETER 
GRADE 1 
MILD 
GRADE 2 
MODERATE 
GRADE 3 
SEVERE 
GRADE 4 
POTENTIALLY 
LIFE-THREATENING 
Unintentional weight 
loss 
NA 
5 – 9% loss in body 
weight from baseline 
10 – 19% loss in body 
weight from baseline 
 20% loss in body 
weight from baseline 
OR  Aggressive 
intervention indicated 
[e.g., tube feeding or 
total parenteral nutrition 
(TPN)] 
INFECTION 
Infection (any other 
than HIV infection) 
Localized, no 
systemic 
antimicrobial 
treatment indicated 
AND Symptoms 
causing no or 
minimal interference 
with usual social & 
functional activities 
Systemic antimicrobial 
treatment indicated 
OR Symptoms 
causing greater than 
minimal interference 
with usual social & 
functional activities 
Systemic antimicrobial 
treatment indicated 
AND Symptoms 
causing inability to 
perform usual social & 
functional activities OR 
Operative intervention 
(other than simple 
incision and drainage) 
indicated 
Life-threatening 
consequences (e.g., 
septic shock) 
INJECTION SITE REACTIONS 
Injection site pain 
(pain without 
touching) 
Or 
Tenderness (pain 
when area is touched)  
Pain/tenderness 
causing no or 
minimal limitation of 
use of limb 
Pain/tenderness 
limiting use of limb 
OR Pain/tenderness 
causing greater than 
minimal interference 
with usual social & 
functional activities 
 
Pain/tenderness 
causing inability to 
perform usual social & 
functional activities 
 
Pain/tenderness 
causing inability to 
perform basic self-care 
function OR 
Hospitalization (other 
than emergency room 
visit) indicated for 
management of 
pain/tenderness 
Injection site reaction (localized) 
Adult > 15 years 
 
Erythema OR 
Induration 
of 5x5 cm – 9x9 cm 
(or 25 cm2 – 81cm2) 
Erythema OR 
Induration OR Edema  
> 9 cm any diameter 
(or > 81 cm2) 
Ulceration OR 
Secondary infection OR 
Phlebitis OR Sterile 
abscess OR Drainage 
Necrosis (involving 
dermis and deeper 
tissue) 
 
Pediatric  15 
years 
Erythema OR 
Induration OR Edema 
present but  2.5 cm 
diameter 
Erythema OR 
Induration OR Edema 
> 2.5 cm diameter but 
< 50% surface area of 
the extremity segment 
(e.g., upper arm/thigh) 
Erythema OR Induration 
OR Edema involving  
 50% surface area of 
the extremity segment 
(e.g., upper arm/thigh) 
OR Ulceration OR 
Secondary infection OR 
Phlebitis OR Sterile 
abscess OR Drainage 
Necrosis (involving 
dermis and deeper 
tissue) 


 
 
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PARAMETER 
GRADE 1 
MILD 
GRADE 2 
MODERATE 
GRADE 3 
SEVERE 
GRADE 4 
POTENTIALLY 
LIFE-THREATENING 
Pruritus associated 
with injection  
See also Skin: 
Pruritus (itching - no 
skin lesions) 
Itching localized to 
injection site AND 
Relieved 
spontaneously or with 
< 48 hours treatment  
Itching beyond the 
injection site but not 
generalized OR 
Itching localized to 
injection site requiring 
 48 hours treatment  
Generalized itching 
causing inability to 
perform usual social & 
functional activities 
NA 
SKIN – DERMATOLOGICAL 
Cutaneous reaction – 
rash 
Localized macular 
rash  
Diffuse macular, 
maculopapular, or 
morbilliform rash OR 
Target lesions  
Diffuse macular, 
maculopapular, or 
morbilliform rash with 
vesicles or limited 
number of bullae OR 
Superficial ulcerations 
of mucous membrane 
limited to one site 
Extensive or 
generalized bullous 
lesions OR Stevens-
Johnson syndrome OR 
Ulceration of mucous 
membrane involving 
two or more distinct 
mucosal sites OR Toxic 
epidermal necrolysis 
(TEN) 
Pruritus (itching – no 
skin lesions)  
(See also Injection 
Site Reactions: 
Pruritus associated 
with injection) 
Itching causing no or 
minimal interference 
with usual social & 
functional activities 
Itching causing 
greater than minimal 
interference with 
usual social & 
functional activities 
Itching causing inability 
to perform usual social 
& functional activities  
NA 
 
CARDIOVASCULAR 
Hypertension 
Adult > 17 years 
(with repeat 
testing at same 
visit) 
140 – 159 mmHg 
systolic 
OR 
90 – 99 mmHg 
diastolic 
160 – 179 mmHg 
systolic 
OR 
100 – 109 mmHg 
diastolic 
≥ 180 mmHg systolic 
OR 
≥ 110 mmHg diastolic 
 
Life-threatening 
consequences (e.g., 
malignant 
hypertension) OR 
Hospitalization 
indicated (other than 
emergency room visit) 
Pediatric  17 
years 
(with repeat 
testing at same 
visit) 
 
NA 
91st –  94th percentile 
adjusted for age, 
height, and gender 
(systolic and/or 
diastolic) 
 
146 - 168 mmHg 
systolic 
 
OR 
 
91 - 102 mmHg 
diastolic 
≥ 95th percentile 
adjusted for age, 
height, and gender 
(systolic and/or 
diastolic) 
 
≥168 mmHg 
systolic 
 
OR 
 
≥102 mmHg 
diastolic 
Life-threatening 
consequences (e.g., 
malignant 
hypertension) OR 
Hospitalization 
indicated (other than 
emergency room visit) 


 
 
Page 86 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
PARAMETER 
GRADE 1 
MILD 
GRADE 2 
MODERATE 
GRADE 3 
SEVERE 
GRADE 4 
POTENTIALLY 
LIFE-THREATENING 
Hypotension 
NA 
Symptomatic, 
corrected with oral 
fluid replacement 
Symptomatic, IV fluids 
indicated 
Shock requiring use of 
vasopressors or 
mechanical assistance 
to maintain blood 
pressure 
 
GASTROINTESTINAL 
Anorexia 
Loss of appetite 
without decreased 
oral intake 
Loss of appetite 
associated with 
decreased oral intake 
without significant 
weight loss 
Loss of appetite 
associated with 
significant weight loss 
Life-threatening 
consequences OR 
Aggressive intervention 
indicated [e.g., tube 
feeding or total 
parenteral nutrition 
(TPN)] 
Comment: Please note that, while the grading scale provided for Unintentional Weight Loss may be used as a guideline when 
grading anorexia, this is not a requirement and should not be used as a substitute for clinical judgment.  
Diarrhea 
Transient or 
intermittent episodes 
of unformed stools 
OR Increase of ≤ 3 
stools over baseline 
per 24-hour period 
Persistent episodes of 
unformed to watery 
stools OR Increase of 
4 – 6 stools over 
baseline per 24-hour 
period 
Bloody diarrhea OR 
Increase of ≥ 7 stools 
per 24-hour period OR 
IV fluid replacement 
indicated 
Life-threatening 
consequences (e.g., 
hypotensive shock) 
Nausea  
Transient (< 24 
hours) or intermittent 
nausea with no or 
minimal interference 
with oral intake 
Persistent nausea 
resulting in decreased 
oral intake for 24 – 48 
hours 
Persistent nausea 
resulting in minimal oral 
intake for > 48 hours 
OR Aggressive 
rehydration indicated 
(e.g., IV fluids) 
Life-threatening 
consequences (e.g., 
hypotensive shock) 
Vomiting 
Transient or 
intermittent vomiting 
with no or minimal 
interference with oral 
intake 
Frequent episodes of 
vomiting with no or 
mild dehydration 
Persistent vomiting 
resulting in orthostatic 
hypotension OR 
Aggressive rehydration 
indicated (e.g., IV 
fluids) 
Life-threatening 
consequences (e.g., 
hypotensive shock) 
Headache  
Symptoms causing 
no or minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
greater than minimal 
interference with 
usual social & 
functional activities 
Symptoms causing 
inability to perform 
usual social & 
functional activities 
Symptoms causing 
inability to perform 
basic self-care functions 
OR Hospitalization 
indicated (other than 
emergency room visit) 
OR Headache with 
significant impairment 
of alertness or other 
neurologic function  
Insomnia 
NA 
Difficulty sleeping 
causing greater than 
minimal interference 
with usual social & 
functional activities 
Difficulty sleeping 
causing inability to 
perform usual social & 
functional activities 
Disabling insomnia 
causing inability to 
perform basic self-care 
functions 


 
 
Page 87 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
PARAMETER 
GRADE 1 
MILD 
GRADE 2 
MODERATE 
GRADE 3 
SEVERE 
GRADE 4 
POTENTIALLY 
LIFE-THREATENING 
RESPIRATORY 
Bronchospasm 
(acute) 
FEV1 or peak flow 
reduced to  
70 – 80% 
FEV1 or peak flow  
50 – 69% 
FEV1 or peak flow  
25 – 49%  
Cyanosis OR FEV1 or 
peak flow < 25% OR 
Intubation 
Dyspnea or respiratory distress 
Adult ≥ 14 years 
Dyspnea on exertion 
with no or minimal 
interference with 
usual social & 
functional activities  
Dyspnea on exertion 
causing greater than 
minimal interference 
with usual social & 
functional activities  
Dyspnea at rest 
causing inability to 
perform usual social & 
functional activities  
Respiratory failure with 
ventilatory support 
indicated 
Pediatric < 14 
years 
Wheezing OR 
minimal increase in 
respiratory rate for 
age 
Nasal flaring OR 
Intercostal retractions 
OR Pulse oximetry 90 
– 95% 
Dyspnea at rest 
causing inability to 
perform usual social & 
functional activities OR 
Pulse oximetry < 90% 
Respiratory failure with 
ventilatory support 
indicated 
MUSCULOSKELETAL 
Arthralgia 
See also Arthritis 
Joint pain causing no 
or minimal 
interference with 
usual social & 
functional activities 
Joint pain causing 
greater than minimal 
interference with 
usual social & 
functional activities 
Joint pain causing 
inability to perform 
usual social & 
functional activities 
Disabling joint pain 
causing inability to 
perform basic self-care 
functions 
Arthritis 
See also Arthralgia 
Stiffness or joint 
swelling causing no 
or minimal 
interference with 
usual social & 
functional activities 
Stiffness or joint 
swelling causing 
greater than minimal 
interference with 
usual social & 
functional activities 
Stiffness or joint 
swelling causing 
inability to perform 
usual social & 
functional activities 
Disabling joint stiffness 
or swelling causing 
inability to perform 
basic self-care functions  
Myalgia 
(non-injection site) 
Muscle pain causing 
no or minimal 
interference with 
usual social & 
functional activities 
Muscle pain causing 
greater than minimal 
interference with 
usual social & 
functional activities 
Muscle pain causing 
inability to perform 
usual social & 
functional activities 
Disabling muscle pain 
causing inability to 
perform basic self-care 
functions 
 
 
 
LABORATORY 
PARAMETER 
GRADE 1 
MILD 
GRADE 2 
MODERATE 
GRADE 3 
SEVERE 
GRADE 4 
POTENTIALLY 
LIFE-THREATENING 


 
 
Page 88 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
Hemoglobin (Hgb) 
 
10.0 – 10.9 g/dL 
6.18 – 6.79 mmol/L 
OR 
Any decrease  
2.5 – 3.4 g/dL 
1.58 – 2.13 mmol/L 
9.0 – 9.9 g/dL 
5.55 - 6.17 mmol/L 
OR 
Any decrease  
3.5 – 4.4 g/dL 
2.14 – 2.78 mmol/L 
7.0 – 8.9 g/dL 
4.34 - 5.54 mmol/L 
OR 
Any decrease  
 4.5 g/dL 
 2.79 mmol/L 
< 7.0 g/dL 
< 4.34 mmol/L 
 
Comment:  The decrease is a decrease from baseline 
Platelets, decreased 
100,000 –  
124,999/mm3 
100.000 x 109 – 
124.999 x 109/L 
50,000 –  
99,999/mm3 
50.000 x 109 – 
99.999 x 109/L 
25,000 –  
49,999/mm3 
25.000 x 109 – 
49.999 x 109/L 
< 25,000/mm3 
< 25.000 x 109/L 
WBC, decreased 
2,000 – 2,500/mm3 
2.000 x 109 –  
2.500 x 109/L 
1,500 – 1,999/mm3 
1.500 x 109 –  
1.999 x 109/L 
1,000 – 1,499/mm3 
1.000 x 109 –  
1.499 x 109/L 
< 1,000/mm3 
< 1.000 x 109/L 
WBC, increased 
 
10,800 – 15,000/mm3 
10.8 – 15.0 x 109/L 
 
15,001 – 20,000/mm3 
15.1 – 20.0 x 109/L 
 
20,001 – 25,000/mm3 
20.1 – 25.0 x 109/L 
 
>25,000/mm3 
>25.0 x 109/L 
 
LABORATORY 
PARAMETER 
GRADE 1 
MILD 
GRADE 2 
MODERATE 
GRADE 3 
SEVERE 
GRADE 4 
POTENTIALLY 
LIFE-THREATENING 
CHEMISTRIES 
 
Standard International Units are listed in italics  
Alkaline Phosphatase 
1.25 – 2.5 x ULN† 
2.6 – 5.0 x ULN† 
5.1 – 10.0 x ULN† 
> 10.0 x ULN† 
ALT (SGPT) 
1.25 – 2.5 x ULN 
2.6 – 5.0 x ULN 
5.1 – 10.0 x ULN 
> 10.0 x ULN 
AST (SGOT) 
1.25 – 2.5 x ULN 
2.6 – 5.0 x ULN 
5.1 – 10.0 x ULN 
> 10.0 x ULN 
Blood urea nitrogen 
(BUN)  
 
 
23 – 26 mg/dL 
8.3 – 9.5 mmol/L 
 
27 – 31 mg/dL 
9.6 – 11.2 mmol/L 
 
>31 mg/dL 
>11.2 mmol/L 
Requires dialysis 
Creatinine – elevated 
 
 
1.5 – 1.7 mg/dL 
121 - 145 umol/L  
 
1.8 – 2.0 mg/dL 
146 – 170 umol/L  
 
2.1 – 2.5 mg/dL 
171 – 208 umol/L  
 
>2.5 mg/dL or requires 
dialysis 
>208 umol/L or 
requires dialysis 
 
Bilirubin (Total) 
1.1 – 1.5 x ULN 
1.6 – 2.5 x ULN 
2.6 – 5.0 x ULN 
> 5.0 x ULN 
URINALYSIS 
 
Standard International Units are listed in italics 
Hematuria (microscopic) 
 5 – 10 cells/mm3 
 
> 10 cells/mm3 
 
Gross, with or without 
clots OR with RBC 
casts 
Transfusion indicated 
Proteinuria, random 
collection 
1 + 
2 – 3 + 
4 + 
NA 
Proteinuria, 24 hour collection 


 
 
Page 89 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
Adult and Pediatric  
 10 years 
200 – 999 mg/24 h 
0.200 – 0.999 g/d 
1,000 – 1,999 mg/24 h 
1.000 – 1.999 g/d 
2,000 – 3,500 mg/24 h 
2.000 – 3.500 g/d 
> 3,500 mg/24 h 
> 3.500 g/d 


 
 
Page 90 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
APPENDIX C 
Adverse Events of Special Interest 
 
Adverse events of special interest (AESIs) are events which are potentially immune mediated 
and include: 
 
Acute disseminated encephalomyelitis (ADEM) 
Addison's Disease 
Anti-neutrophil Cytoplasmic Antibody (ANCA)-associated Vasculitis 
Ankylosing Spondylitis 
Anti-phospholipid Syndrome 
Autoimmune Bullous Skin Diseases 
Autoimmune Hemolytic Anemia 
Autoimmune Hepatitis 
Autoimmune Thrombotic/Thromboembolic Conditions 
Basedow’s Disease 
Behcet’s Syndrome 
Bell's Palsy 
Carditis 
Celiac Disease 
Crohn’s Disease 
Cutaneous Lupus 
Demyelinating Disease 
Dermatomyositis 
Diabetes Mellitus, Insulin Dependent (IDDM) 
Erythema Nodosum 
Glomerulonephritis 
Guillain Barre Syndrome 
Grave’s Disease 
Idiopathic Thrombocytopenic Purpura (ITP) 
Inflammatory Bowel Disease (non-specific) 
Juvenile Rheumatoid Arthritis 
Mixed Connective Tissue Disease 
Multiple Sclerosis 
Myasthenia Gravis 
Myelitis/Transverse Myelitis 
Myocarditis 
Nephritis 
Optic neuritis 
Pericarditis 
Polymyalgia Rheumatica 
Polymyositis 
Primary Biliary Cirrhosis 
Primary Sclerosing Cholangitis 
Psoriasis 


 
 
Page 91 of 91 
 
 
AERAS-404 
 
Protocol C-040-404  
 
CONFIDENTIAL 
25/Jul/2016; Final Version 5.0 
 
Aeras Clinical Protocol Template Edition 01.5; Reference SOP CLD-136 rev. 01, ATT-CLD136-B 
Psoriatic Arthritis 
Raynaud's Phenomenon 
Rheumatoid Arthritis 
Sarcoidosis 
Scleroderma 
Sjogren’s Syndrome 
Spondylo-arthropathy 
Stevens-Johnson Syndrome 
Systemic Lupus Erythematosus 
Temporal Arteritis 
Thyroiditis 
Tolosa-Hunt Syndrome  
Ulcerative Colitis 
Ulcerative Proctitis 
Uveitis 
Vasculitis 
Vitiligo 
Wegener’s Granulomatosis 

SAP_001.pdf: 
 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
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disclosure or reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 1 of 91 
STATISTICAL ANALYSIS PLAN 
FINAL ANALYSIS 
 
 
C-040-404 
 
 
A RANDOMIZED, PLACEBO CONTROLLED, PARTIALLY BLINDED PHASE II 
STUDY TO EVALUATE SAFETY, IMMUNOGENICITY, AND PREVENTION OF 
INFECTION WITH MYCOBACTERIUM TUBERCULOSIS OF AERAS-404 AND 
BCG REVACCINATION IN HEALTHY ADOLESCENTS 
 
 
AUTHOR: LOUISE VAN ASWEGEN 
VERSION NUMBER AND DATE: V2.0, 29SEP2017 
 
 


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
The contents of this document are confidential and proprietary to Quintiles Transnational Corp. Unauthorized use, disclosure or 
reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 2 of 91 
STATISTICAL ANALYSIS PLAN SIGNATURE PAGE 
Statistical Analysis Plan Version 2.0 (Dated 29SEP2017) for Protocol C-040-404. 
 
Name 
Signature 
Date (DDMMMYYY) 
Author: 
Louise van Aswegen 
 
 
 
Position: 
Statistical Team Lead 
 
Company: 
QuintilesIMS 
 
Upon review of this document, the undersigned approve this version of the Statistical Analysis Plan, authorizing 
that the content is acceptable for the final analysis. 
 
Name 
Signature 
Date (DDMMMYYYY) 
Approved by: 
Kathryn Rutkowski 
 
 
 
Position: 
Director, Biostatistics and Data Management 
 
Company: 
Aeras 
 
Approved by: 
Robert Hopkins 
 
 
 
Position: 
Vice President, Product Development 
 
Company: 
Aeras 
 
Approved by: 
Ann Ginsberg 
 
 
Position: 
Chief Medical Officer 
Company: 
Aeras 
 
Approved by: 
Mark Riddle 
 
 
Position: 
Global Safety Representative  
Company: 
Aeras 
 
 


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
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reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 3 of 91 
 
Name 
Signature 
Date (DDMMMYYYY) 
Approved by: 
Danilo Casimiro 
 
 
Position: 
Chief Scientific Officer   
Company: 
Aeras 
 
Approved by: 
Barbara Shepherd 
 
Position: 
Director, Medical Writing   
Company: 
Aeras 
 
 


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
The contents of this document are confidential and proprietary to Quintiles Transnational Corp. Unauthorized use, disclosure or 
reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 4 of 91 
MODIFICATION HISTORY 
 
Unique 
Identifier for 
this Version 
Date of the 
Document 
Version 
Author 
Significant Changes from  
Previous Authorized Version 
1.0 
26JUL2017 
Louise van Aswegen 
Not Applicable. First Version. 
2.0 
29SEP2017 
Louise van Aswegen 
Update alternative threshold text from 
‘<0.2 at Baseline and > 0.70 post-Day 
84’ to ‘<0.20 at any time point prior to 
conversion and > 0.70 at any time point 
post-Day 84’ 
Repeat the secondary analyses for this 
threshold.  
Add analyses of whole blood ICS to be 
performed by Aeras 
 
 
 


 
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Template No:  CS_TP_BS016 – Revision 3 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 5 of 91 
TABLE OF CONTENTS 
1. INTRODUCTION ...........................................................................................................................10 
2. STUDY OBJECTIVES ....................................................................................................................11 
2.1. 
Primary Objectives ................................................................................................................................. 11 
2.2. 
Secondary Objectives ............................................................................................................................. 11 
2.3. 
Exploratory Objectives ........................................................................................................................... 11 
3. STUDY DESIGN .............................................................................................................................12 
3.1. 
General Description ................................................................................................................................ 12 
3.2. 
Schedule of Events.................................................................................................................................. 14 
3.3. 
Changes to Analysis from Protocol ......................................................................................................... 16 
4. PLANNED ANALYSES ..................................................................................................................18 
4.1. 
Data Monitoring Committee (DMC) ........................................................................................................ 18 
4.2. 
Primary Analysis ..................................................................................................................................... 18 
4.3. 
Final Analysis .......................................................................................................................................... 19 
5. ANALYSIS SETS ............................................................................................................................19 
5.1. 
Randomized Analysis Set ........................................................................................................................ 19 
5.2. 
Safety Analysis Set .................................................................................................................................. 19 
5.3. 
Modified Intent-to-treat Analysis Set ..................................................................................................... 20 
5.4. 
Adjusted Modified Intent-to-treat (first recall positive omit Analysis Set) .............................................. 21 
5.5. 
Per Protocol Analysis Set ........................................................................................................................ 21 
6. GENERAL CONSIDERATIONS ...................................................................................................23 
6.1. 
Reference Start Date and Study Day ....................................................................................................... 23 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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6.2. 
Baseline .................................................................................................................................................. 23 
6.3. 
Retests, Unscheduled Visits and Premature Discontinuation Data ......................................................... 23 
6.4. 
Statistical Tests ....................................................................................................................................... 24 
6.5. 
Common Calculations ............................................................................................................................. 24 
6.6. 
Software Version .................................................................................................................................... 25 
7. STATISTICAL CONSIDERATIONS ...........................................................................................25 
7.1. 
Adjustments for Covariates and Factors to be Included in Analyses ....................................................... 25 
7.2. 
Multicenter Studies ................................................................................................................................ 25 
7.3. 
Missing data ........................................................................................................................................... 25 
7.4. 
Multiple Comparisons/Multiplicity ......................................................................................................... 25 
7.5. 
Examination of Subgroups ...................................................................................................................... 26 
8. OUTPUT PRESENTATIONS .......................................................................................................26 
9. DISPOSITION AND PREMATURE DISCONTINUATION ....................................................26 
9.1. 
Derivations ............................................................................................................................................. 29 
10. 
DEMOGRAPHIC AND OTHER BASELINE CHARACTERISTICS ....................................29 
10.1. 
Derivations ......................................................................................................................................... 31 
11. 
MEDICAL HISTORY .................................................................................................................31 
12. 
PRIOR AND CONCOMITANT MEDICATIONS ...................................................................32 
13. 
STUDY VACCINATION ............................................................................................................32 
13.1. 
Derivations ......................................................................................................................................... 33 
14. 
EFFICACY OUTCOMES ............................................................................................................34 
14.1. 
Primary Efficacy .................................................................................................................................. 34 


 
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14.1.1. 
Primary Efficacy Endpoint ...................................................................................................................... 34 
14.1.2. 
Analysis of Primary Efficacy Endpoint .................................................................................................... 34 
14.1.3. 
Derivations ............................................................................................................................................. 37 
14.2. 
Secondary Efficacy .............................................................................................................................. 40 
14.2.1. 
Secondary Efficacy Endpoint ................................................................................................................. 40 
14.2.2. 
Analysis of Secondary Efficacy Endpoint ............................................................................................... 40 
14.2.3. 
Derivations ............................................................................................................................................. 41 
14.3. 
Exploratory Efficacy ............................................................................................................................ 42 
14.3.1. 
Exploratory Efficacy Endpoint ................................................................................................................ 42 
14.3.2. 
Analysis of Exploratory Efficacy Endpoint .............................................................................................. 43 
14.3.3. 
Derivations ............................................................................................................................................. 47 
15. 
IMMUNOGENICITY ..................................................................................................................51 
15.1. 
Peripheral Blood Mononuclear Cells (PBMC) Intracellular Cytokine Staining (ICS) .............................. 51 
15.1.1. 
Derivations ............................................................................................................................................. 54 
15.1.1.1. 
Negative control-subtracted Cytokine Response .......................................................................... 54 
15.1.1.2. 
Single Positive CD107a ................................................................................................................... 54 
15.1.1.3. 
Total Number of Cells .................................................................................................................... 54 
15.1.1.4. 
ANY Response ................................................................................................................................ 54 
15.1.1.5. 
Total Stimulation Antigen Result ................................................................................................... 54 
15.1.1.6. 
Positive (G,2) .................................................................................................................................. 55 
15.1.2. 
Responder Status ................................................................................................................................... 55 
15.2. 
Whole blood ICS ................................................................................................................................. 57 
15.2.1. 
Derivations ............................................................................................................................................. 58 
15.2.1.1. 
Positive (G,2,T,17,22) ..................................................................................................................... 58 
15.2.1.2. 
Positive (G,2,T) ............................................................................................................................... 58 
15.2.1.3. 
Positive (17) ................................................................................................................................... 58 
15.2.1.4. 
Positive (22) ................................................................................................................................... 58 
15.2.2. 
Responder Status ................................................................................................................................... 59 
16. 
SAFETY OUTCOMES ................................................................................................................59 
16.1. 
Adverse Events (AEs) .......................................................................................................................... 59 
16.1.1. 
Derivations ............................................................................................................................................. 60 
16.1.2. 
All Adverse Events (AEs) ........................................................................................................................ 60 
16.1.2.1. 
Severity .......................................................................................................................................... 61 
16.1.2.2. 
Relationship to Study Vaccination ................................................................................................. 62 
16.1.3. 
Unsolicited Adverse Events (AEs) .......................................................................................................... 63 
16.1.4. 
Solicited and Injection Site Adverse Events (AEs) .................................................................................. 63 
16.1.5. 
Serious Adverse Events (SAEs) ............................................................................................................... 64 
16.1.6. 
Adverse Events of Special Interest (AESI) .............................................................................................. 64 
16.1.7. 
Adverse Events Leading to Premature Study Discontinuation .............................................................. 64 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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16.1.8. 
Adverse Event (AE) Dose at Onset (Most Recent Vaccination) ............................................................. 65 
16.1.9. 
Adverse Event (AE) Post-vaccination Reporting Window ...................................................................... 65 
16.2. 
Injection Site Reaction Assessment ..................................................................................................... 66 
16.2.1. 
Derivations ............................................................................................................................................. 67 
16.3. 
Laboratory Evaluations ....................................................................................................................... 67 
16.3.1. 
Toxicity Grading Criteria ........................................................................................................................ 68 
16.4. 
Vital Signs ........................................................................................................................................... 68 
16.4.1. 
Toxicity Grading Criteria ........................................................................................................................ 69 
16.4.2. 
Pre-Vaccination and Post-Vaccination Time Points ............................................................................... 69 
16.5. 
Physical Examination .......................................................................................................................... 69 
16.6. 
Pregnancies ........................................................................................................................................ 70 
16.6.1. 
Derivations ............................................................................................................................................. 70 
17. 
DATA NOT SUMMARIZED OR PRESENTED .....................................................................71 
18. 
REFERENCES .............................................................................................................................72 
APPENDIX 1. PROGRAMMING CONVENTIONS FOR OUTPUTS .............................................73 
Study Visits ........................................................................................................................................................ 73 
Footnotes ........................................................................................................................................................... 73 
Output Headers ................................................................................................................................................. 73 
Output Footers .................................................................................................................................................. 73 
Decimal Precision ............................................................................................................................................... 73 
Dates & Times .................................................................................................................................................... 74 
Spelling Format .................................................................................................................................................. 74 
Listings ............................................................................................................................................................... 74 
Presentation Notes ............................................................................................................................................ 74 
APPENDIX 2. PARTIAL DATE CONVENTIONS ............................................................................76 


 
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APPENDIX 3. SOLICITED AND INJECTION SITE ADVERSE EVENT (AE) PREFERRED 
TERMS AND CODES ............................................................................................................................78 
APPENDIX 4. ADVERSE EVENTS OF SPECIAL INTEREST (AESIS) ........................................79 
APPENDIX 5. LABORATORY ASSESSMENTS ...............................................................................80 
APPENDIX 6. VITAL SIGNS ASSESSMENTS ..................................................................................81 
APPENDIX 7. TOXICITY GRADING CRITERIA .............................................................................82 
Estimating Severity Grade .................................................................................................................................. 82 
Cardiovascular ................................................................................................................................................... 82 
Skin – Dermatological ........................................................................................................................................ 83 
Infection ............................................................................................................................................................ 83 
Systemic ............................................................................................................................................................. 84 
Injection Site Reactions ...................................................................................................................................... 85 
Gastrointestinal ................................................................................................................................................. 86 
Respiratory ........................................................................................................................................................ 87 
Musculoskeletal ................................................................................................................................................. 88 
Laboratory: Hematology .................................................................................................................................... 89 
Laboratory: Chemistry ....................................................................................................................................... 89 
Laboratory: Urinalysis ........................................................................................................................................ 90 
APPENDIX 8. PROTOCOL-DEFINED VISIT WINDOWS .............................................................91 
 
 


 
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1. 
INTRODUCTION 
This document describes the rules and conventions that will be used in the planned presentation and analysis 
of data of the final clinical study report (CSR) for Protocol C-040-404 as set out in the relevant version of the 
table, listing and figure (TLF) shells. It describes the data to be summarized and analyzed, including specifics of 
the statistical analyses to be performed on the data.  
This SAP is based on protocol Final Version 5.0, dated 25JUL2016. 
 
 


 
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2. 
STUDY OBJECTIVES 
The following relates to the overall study objectives as described in Section 2.0: Study Objectives and Design of 
the protocol. 
2.1. PRIMARY OBJECTIVES 
The primary objectives are: 
• 
To evaluate the safety profile in human immunodeficiency virus (HIV)-uninfected, remotely Bacillus 
Calmette-Guérin (BCG) vaccinated adolescents of: 
o 
H4:IC31 (also known as AERAS-404). 
o 
BCG revaccination. 
• 
To evaluate prevention of Mycobacterium tuberculosis (Mtb) infection, as measured by rates of 
conversion using a QuantiFERON Tuberculosis (TB) gold-in-tube (QFT-GIT) assay of: 
o 
H4:IC31 compared to Placebo. 
o 
BCG revaccination compared to Placebo. 
2.2. SECONDARY OBJECTIVES 
The secondary objectives are: 
• 
To evaluate prevention of Mtb infection, as measured by rates of sustained conversion using a QFT-GIT 
assay of: 
o 
H4:IC31 compared to Placebo. 
o 
BCG revaccination compared to Placebo. 
• 
To investigate the immunogenicity in HIV-uninfected, remotely BCG vaccinated adolescents of: 
o 
H4:IC31. 
o 
BCG revaccination. 
2.3. EXPLORATORY OBJECTIVES 
The exploratory objectives are: 
• 
To evaluate prevention of Mtb infection, as measured by rates of reversion to a negative from a positive 
QFT-GIT assay of: 
o 
H4:IC31 compared to Placebo. 
o 
BCG revaccination compared to Placebo. 
• 
To explore the effect of alternative QFT-GIT test threshold values on rates of QFT-GIT conversion, QFT-
GIT reversion and prevention of Mtb infection. 
• 
To identify immune correlates of risk for Mtb infection. 


 
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• 
To identify immune correlates of vaccine-induced protection against Mtb infection induced by H4:IC31 or 
BCG revaccination. 
• 
To evaluate alternative interferon gamma release assays (IGRAs) or immune markers for diagnosis of Mtb 
infection or as markers of exposure to nontuberculous mycobacteria (NTM). 
• 
To explore trends in TB disease incidence after H4:IC31 vaccination or BCG revaccination. 
• 
To explore trends in QFT-GIT prolonged/sustained conversions and late reversions (i.e., more than 6 
months post initial conversion) in early QFT-GIT converters (i.e., among those who converted at Month 6 
or Month 12 of follow-up). 
3. 
STUDY DESIGN 
3.1. GENERAL DESCRIPTION 
A Phase II, randomized, 3-arm, placebo-controlled, partially blinded clinical trial that was conducted in 990 
healthy HIV-uninfected, QFT-GIT negative, previously BCG vaccinated adolescents. Participants were recruited 
from high schools or directly from the community. 
Participants were enrolled in two sequential cohorts: 
• 
Safety and Immunogenicity Cohort: 
o 
First 90 participants (30 from each arm). 
o 
More intensive collection of safety data. 
o 
Selected immunogenicity assays (including whole blood intracellular cytokine staining [ICS]). 
o 
Data reviewed by the data monitoring committee (DMC), principal investigator and local medical 
monitor. 
• 
Correlates Cohort: 
o 
Remaining 900 participants. 
All 990 participants in the study were evaluated for: 
• 
Safety outcomes. 
• 
Biomarker outcomes. 
• 
Prevention of Mtb infection. 
Within each cohort, participants were randomized in a 1:1:1 ratio to one of the following treatment groups: 
• 
H4:IC31: 
o 
15 mcg H4/500 nmol IC31. 
o 
Two doses (administered on Day 0 and Day 56). 
o 
Intramuscular injection. 
• 
Placebo: 
o 
Saline volume equivalent to the H4:IC31 injection. 
o 
Two doses (administered on Day 0 and Day 56). 
o 
Intramuscular injection. 


 
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• 
BCG: 
o 
2-8 x 105 CFU. 
o 
One dose (administered on Day 0). 
o 
Intradermal injection. 
Randomization to the H4:IC31 and Placebo treatment groups was double-blinded. Since BCG causes a 
recognizable local injection site reaction, the BCG revaccination treatment group was not blind. Laboratory 
staff performing QFT-GIT, immunogenicity and correlates assays were blinded to all three treatment groups. 
Planned randomization of participants is specified in Table 1: Planned Enrollment by Treatment Group and 
Cohort, below: 
 Table 1: Planned Enrollment by Treatment Group and Cohort 
Cohort 
Planned Number of Participants 
Treatment Group 
Total 
Placebo 
BCG 
H4:IC31 
Safety and Immunogenicity 
30 
30 
30 
90 
Correlates 
300 
300 
300 
900 
Total 
330 
330 
330 
990 
BCG: Bacillus Calmette-Guerin. 
Safety outcomes included solicited and unsolicited adverse events (AEs), injection site assessment, adverse 
events of special interest (AESIs) and serious adverse events (SAEs). 
Each participant was followed for safety for a minimum of 6 months after the last vaccination. 
The primary endpoint is: 
• 
QFT-GIT conversion from negative to positive at any time point after Day 84 through end of follow-up in 
participants who were QFT-GIT negative at Study Day 84. 
The secondary endpoint is: 
• 
The primary endpoint occurrence (see aforementioned), persisting without QFT-GIT reversion from 
positive to negative through 6 months after QFT-GIT conversion. 
The duration of the trial to primary analysis was endpoint-driven. The primary analysis was triggered when all 
of the following conditions were met: 
• 
At least 64 primary endpoints were accrued in the study (regardless from which cohort). 
• 
The median follow-up time was at least 15 months: 
o 
Maximum individual follow-up time to detect QFT-GIT conversion of 24 months. 
The protocol stated that if an initial reduction in infection rate in the BCG or H4:IC31 treatment group was seen 
in the primary analysis, follow-up for the entire study may be extended to examine the duration of prevention 
of Mtb infection. Following the primary analysis, there were no safety concerns of note.  The DMC 
recommended that follow-up be continued as per protocol with a further DMC review close to the end of 
follow-up.  Aeras confirmed that this second review by the DMC will only be completed following database (DB) 
lock. They also recommended that all participants have a QuantiFERON test performed at 24 months, including 


 
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all converters. The Sanofi Pasteur-Aeras Joint Steering Committee convened on June 14, 2016 and agreed to 
follow the DMC recommendations and to maintain study blind until study completion.  
All participants with a primary endpoint (QFT-GIT conversion) were followed for an additional 6 months post-
conversion to confirm whether they met the secondary or exploratory endpoints of sustained conversion or 
QFT reversion. 
Participants who converted to a positive QFT-GIT at Month 6 or Month 12 were approached for a final QFT-GIT 
test and assessment for TB signs and symptoms at least 24 months after their initial vaccination. 
The schedule for follow-up for individual participants was contingent on results of the QFT-GIT tests conducted 
at Month 3 (Day 84) and Months 6, 12, 18 and 24 study visits. A schematic of follow-up and QFT-GIT testing is 
presented in Figure 1: Schematic of Follow-up and QFT-GIT Testing, below. 
 
Figure 1: Schematic of Follow-up and QFT-GIT Testing 
*Participants who converted at Month 6 or Month 12 were asked to return for a repeated QFT-GIT test done 
at least 24 months after their initial vaccination.  
3.2. SCHEDULE OF EVENTS 
The schedules of events for the BCG participants as well as the H4:IC31/Placebo participants are presented in 
Table 2: Schedule of Participant Evaluations (Participants Receiving BCG, N = 330) and Table 3: Schedule of 
Participant Evaluations (Participants Receiving H4:IC31 [N=330] or Placebo [N=330]), below. 
 
 


 
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Table 2: Schedule of Participant Evaluations (Participants Receiving BCG, N = 330) 
Study Visit Day (D) or Month (M)  
Screen 
0 
D7 
D28 
D70 
D84c 
M6 (D168)d,e 
M12 (D336) 
M18 (D504) 
M24 (D672) 
Eligibility criteria verification 
X 
X 
 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
Physical examination 
X 
 
 
 
 
 
 
Urine HCG (all females) 
X 
X 
 
 
 
 
 
QuantiFERON-TB Gold in-tube (mL) 
3 
 
 
 
 
3 
3 
HIV-1 (mL) with HIV counselling  
3 
 
 
 
 
 
 
Urinalysis 
X 
 
Xb 
 
 
 
 
Serum chemistry (mL)g 
5 
 
5b 
 
 
 
 
CBC, differential, platelets (mL) 
5 
 
5b 
 
 
 
 
Vital signs 
X 
X 
 
 
 
 
 
Interval history 
 
X 
X 
X 
X 
X 
X 
Focused physical examinationa 
 
X 
X 
X 
X 
X 
X 
Distribute/review diary cards 
 
Xb 
Xb 
 
 
 
 
Solicited adverse events (incl. con. meds.) 
 
X 
X 
 
 
 
 
Unsolicited adverse events (incl. con. meds.) 
 
X 
X 
X 
 
 
 
SAEs, AESIs, and SUSARs (incl. con. meds.) 
 
X 
X 
X 
X 
X 
X 
Solicited and unsolicited injection site reaction adverse events (incl. con. meds.)  
 
X 
X 
X 
X 
X 
 
Site of injection examination 
 
X 
X 
X 
X 
X 
 
TB symptom screen 
X 
 
 
X 
X 
X 
X 
BCG administration 
 
X 
 
 
 
 
 
Whole blood assay (mL) 
 
6b 
 
 
6b 
 
 
Absolute blood count (mL) 
 
0.
5 
0.
5 
 
 
 
0.5 
RNA (mL) 
 
2.
5 
2.
5 
 
 
 
2.5 
PBMC for ICS  (mL) 
 
8b 
 
 
8b 
 
 
Serum for multiplex ELISA (mL) 
 
3b 
3b 
 
 
 
 
PBMC, plasma, and serum for correlates of risk/protection (mL) 
 
1
6 
 
 
16 
 
40i 
Whole blood for evaluation of novel IGRA (mL)f 
7 
7 
 
 
 
7 
7 
PBMC for evaluation of NTM exposure (mL) 
 
1
0 
 
 
 
 
 
Per visit phlebotomy volume (mL)h 
23 
5
3 
16 
0 
32.5 
10 
53 (M6,M12)/ 
13 (M18,M24) 
Cumulative phlebotomy volume (mL)h 
23 
7
6 
92 
92 
124.5 
134.5 
266.5  
 
a. 
If indicated by interval history 
b. 
Subjects in the Safety & Immunogenicity Cohort only 
c. 
Subjects who convert from QFT-GIT(-) to (+) at Day 84 will have a final visit at 168 days ±14 days after vaccination for assessment of SAEs, AESIs, SUSARs, and TB symptom screen 
d. 
Month 6, 12, 18, and 24 study visits will be completed by subjects who did not convert from QFT-GIT (-) to (+) at the previous visit; the Month 24 visit will be the final visit for subjects 
who remain QFT-GIT(-) throughout the study.  
e. 
Subjects who convert from QFT-GIT (-) to (+) at Month 6-24 visits will have two additional visits with repeat QFT-GIT testing, assessment of SAEs, AESIs, SUSARs, and TB symptom 
screen 84 days ±14 days and 168 days ±14 days after conversion. Up to 10 mL of blood will be collected at each of those visits, including blood for absolute blood count and RNA. 
Subjects who initially convert to QFT-GIT(+) at Month 6 or 12 will be asked to have a final QFT-GIT test, blood collection for absolute blood count and RNA, and TB symptom screen 
done at least 24 months after their initial vaccination.  
f. 
May not be collected at all time points shown. 
g. 
Serum chemistry includes AST, ALT, alkaline phosphatase, total bilirubin, creatinine, and BUN. 
h. 
Blood volumes are approximate. 
i. 
Study Months 6 and 12 only. 
Table 3: Schedule of Participant Evaluations (Participants Receiving H4:IC31 [N=330] or Placebo [N=330]) 
Study Visit Day  
(D) or Month (M)  
Screen 
0 
D3  
D7 
D28 
D56 
D63 
D70 
D84c  
M6  (D168)d,e 
M12 (D336) 
M18 (D504) 
M24 (D672) 
Eligibility criteria verification 
X 
X 
 
 
 
X 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
 
 
 
Physical examination 
X 
 
 
 
 
 
 
 
 
 
Urine HCG (all females) 
X 
X 
 
 
 
X 
 
 
 
 
QuantiFERON-TB Gold in-tube (mL) 
3 
 
 
 
 
 
 
 
3 
3 
HIV-1 (mL) with HIV counselling  
3 
 
 
 
 
 
 
 
 
 
Urinalysis 
X 
 
 
Xb 
 
 
Xb 
 
 
 
Serum chemistry (mL)g 
5 
 
 
5b 
 
 
5b 
 
 
 
CBC, differential, platelets (mL) 
5 
 
 
5b 
 
 
5b 
 
 
 
Vital signs 
X 
X 
 
 
 
X 
 
 
 
 


 
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Study Visit Day  
(D) or Month (M)  
Screen 
0 
D3  
D7 
D28 
D56 
D63 
D70 
D84c  
M6  (D168)d,e 
M12 (D336) 
M18 (D504) 
M24 (D672) 
Interval history 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Focused physical examinationa 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Distribute/review diary cards 
 
Xb 
 
Xb 
 
Xb 
Xb 
 
 
 
Solicited adverse events (incl. con. meds.) 
 
X 
X 
X 
 
X 
X 
 
 
 
Unsolicited adverse events (incl. con. meds.) 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
SAEs, AESIs, and SUSARs (incl. con. meds.) 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Solicited injection site reaction adverse events 
(incl. con. meds.)  
 
X 
X 
X 
X 
X 
X 
X 
X 
 
Site of injection examination 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
TB symptom screen 
X 
 
 
 
X 
X 
 
 
X 
X 
Study vaccine administration 
 
X 
 
 
 
X 
 
 
 
 
Whole blood assay (mL) 
 
 6b  
 
 
 
 
 
6b 
 
 
Absolute blood count (mL) 
 
0.
5 
0.
5 
 
 
 
 
 
 
0.5 
RNA (mL) 
 
2.
5 
2.
5 
 
 
 
 
 
 
2.5 
PBMC for ICS (mL) 
 
8b 
 
 
 
 
 
8b 
 
 
Serum for multiplex ELISA (mL) 
 
3b 
3b 
 
 
 
 
 
 
 
PBMC, plasma, and serum for correlates of 
risk/protection (mL) 
 
16 
 
 
 
 
 
16 
 
40i 
Whole blood for evaluation of novel IGRA (mL)f 
7 
7 
 
 
 
 
 
 
7 
7 
PBMC for evaluation of  NTM exposure (mL) 
 
10 
 
 
 
 
 
 
 
 
Per visit phlebotomy volume (mL)h 
23 
53 
6 
10 
0 
0 
10 
32.5 
10 
53 (M6,M12)/ 
13 (M18,M24) 
Cumulative phlebotomy volume (mL)h 
23 
76 
82 
92 
92 
92 
102 
134.5 
144.5 
276.5  
a. 
If indicated by interval history 
b. 
Subjects in the Safety & Immunogenicity Cohort only 
c. 
Subjects who convert from QFT-GIT(-) to (+) at Day 84 will have a final visit at 168 days ±14 days after second vaccination (Month 8, Study Day 224) for evaluation of SAEs, AESIs, 
SUSARs, and TB symptom screen 
d. 
The Month 6, 12, 18, and 24 study visits will be completed by subjects who did not convert from QFT-GIT (-) to (+) at the previous visit; the Month 24 visit will be the final visit for 
subjects who remain QFT-GIT(-) throughout the study 
e. 
Subjects who convert from QFT-GIT (-) to (+) at Month 6-24 visits will have two additional visits with repeat QFT-GIT testing, assessment of SAEs, AESIs, SUSARs, and TB symptom screen 
84 days ±14 days and 168 days ±14 days after conversion. Up to 10 mL of blood will be collected at each of those visits, including blood for absolute blood count and RNA. Subjects who 
initially convert to QFT-GIT(+) at Month 6 or 12 will be asked to have a final QFT-GIT test, blood collection for absolute blood count and RNA,  and TB symptom screen done at least 24 
months after initial vaccination.   
f. 
May not be collected at all time points shown. 
g. 
Serum chemistry includes AST, ALT, alkaline phosphatase, total bilirubin, creatinine, and BUN. 
h. 
Blood volumes are approximate. 
i. 
Study Months 6 and 12 only. 
 
3.3. CHANGES TO ANALYSIS FROM PROTOCOL 
• 
On page 23 of the protocol, the following is stated: “The secondary analysis will be triggered after 6 
months of additional post-conversion follow-up for individuals who convert, regardless of the rate of 
QFT-GIT reversion.” 
Following discussions with Aeras, it was clarified that there will not be any formal secondary analysis. Following 
the primary analysis there will only be one additional analysis, the final analysis. The final analysis will occur 
once all planned follow-up visits have been completed, or following the termination of the trial (if terminated 
before the required follow-up visits occurred). 
• 
On page 52 of the protocol the Intent-to-treat analysis set is defined as: “The intent-to-treat population 
for efficacy and immunogenicity analyses will consist of all randomized subjects.” 
Following discussions with Aeras, it was decided that since the Intent-to-treat analysis set is the same as the 
Randomized analysis set, a Modified Intent-to-treat analysis set will be defined and used for analysis purposes. 
The Modified Intent-to-treat analysis set is defined as described in Section 5.3: Modified Intent-to-treat 
Analysis Set. 


 
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• 
The following exploratory analyses will not be performed as part of this SAP, as agreed by Aeras: 
o 
Identifying immune correlates of risk for Mtb infection. 
o 
Evaluating alternative IGRAs or immune markers for diagnosis of Mtb infection or as markers of 
exposure to nontuberculous mycobacteria (NTM). 
o 
Identifying immune correlates of vaccine-induced protection against Mtb infection induced by 
H4:IC31 or BCG revaccination. 
o 
Exploring trends in TB disease incidence after vaccination.  
Aeras and some of its external partners will prepare a separate correlates SAP once the trial’s efficacy results 
become available.  
• 
On 5 April 2016, Aeras Biostatistics and Data Management were notified of a manufacturer’s recall of a 
portion of the QuantiFERON (QFT) tubes that were used in determination of endpoints. The recall 
primarily stemmed from changes in Qiagen’s quality control (QC) process, which resulted in their 
recalling all unexpired partial lots that had been previously circulated for use. Although Qiagen’s ongoing 
QC processes did not detect a spike in false positive results within this partial lot, they were unable to 
rule out the possibility of false positive results coming from the tubes in question, thereby resulting in 
the recall.  
o 
Sixteen (16) positive QFT results came from these recalled tubes, affecting 14 unique participants. 
Nine (9) of the tubes contributed toward the trial’s primary efficacy endpoint. Even though the 
information provided by Qiagen does not suggest that these results are false positives, the tubes 
were recalled nonetheless. Therefore, in an abundance of caution, Aeras considered an additional 
analysis set that would remove these results from the primary endpoint and final endpoint analysis.  
Aeras requested the following approach to handle this: 
o 
Create an additional analysis set (refer to Section 5.4: Adjusted Modified Intent-to-treat (first recall 
positive omit Analysis Set)).  
o 
Perform the following additional analysis: 
▪ 
Modified Intent-to-treat (mITT) with adjudication: This analysis will retain QFT data from all visits 
for participants who received positive QFT results from recalled tubes provided they also 
received positive results from non-recalled tubes. In other words, if a participant has positive QFT 
results from both non-recalled and recalled tubes, then all of these results will be treated as true 
positives. 
• 
Whilst setting up the programming for the medical review listings, Data Management (DM) was 
requested to use the following toxicity criteria for Urine Erythrocytes since this is recorded as a 
qualitative value and the protocol provides the toxicity criteria for quantitative values.  These criteria will 
be used to determine the toxicity grading. ++/+++ values will therefore be regarded as ‘Grade 2/3/4’.  
Laboratory Test 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Urine Erythrocytes 
+ 
++/+++ 
++/+++ 
++/+++ 


 
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4. 
PLANNED ANALYSES 
The following formal analyses will be performed for this study: 
• 
Day 7 DMC. 
• 
Day 84 DMC. 
• 
Primary analysis. 
• 
Final analysis. 
4.1. DATA MONITORING COMMITTEE (DMC) 
The DMC safety reviews, based on a separate statistical analysis plan (SAP) and set of TLF shells (as provided by 
Aeras), have been completed at two time points: 
• 
Day 7 DMC: 
o 
For the Safety and Immunogenicity Cohort once the last participant reached Day 7, the DMC 
reviewed unblinded safety data. 
• 
Day 84 DMC: 
o 
For the Safety and Immunogenicity Cohort once the last participant reached Day 84, the DMC 
reviewed unblinded safety data. 
The DMC made a formal recommendation about the continued conduct of the trial after each safety review. 
Enrollment continued while the reviews were being conducted. 
4.2. PRIMARY ANALYSIS 
The primary analysis, based on a separate SAP and set of TLF shells was completed once the following two 
conditions were met: 
• 
At least 64 primary endpoints had accrued (refer to Section 14.1.1: Primary Efficacy Endpoint for a 
description of the primary endpoints). 
• 
The median follow-up time (for all participants in the Modified Intent-to-treat [mITT] analysis set) from 
Day 0 was at least 15 months. 
The DMC received the primary analysis report and provided a recommendation to the study sponsor regarding 
whether to continue, stop or modify the trial as currently planned. 


 
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4.3. FINAL ANALYSIS 
The final analysis identified in this SAP and relevant set of TLF shells will be performed by QuintilesIMS 
Bloemfontein, South Africa (QBLM) Biostatistics. The final analysis is planned following: 
• 
Aeras’s authorization of the final version of the SAP and relevant set of TLF shells. 
• 
Aeras’s authorization of the final analysis sets. 
• 
Final database lock and unblinding.  
The final analysis will be performed on a clean locked database: 
• 
All outstanding data issues and queries resolved. 
• 
All unresolvable data issues documented in a separate data handling report (DHR) document from Data 
Management.  
• 
All coding of medications, medical conditions, physical examination abnormalities and adverse events 
(AEs) completed. 
• 
All reconciliation of vendor data with electronic case report form (eCRF) data completed successfully.  
5. 
ANALYSIS SETS 
Aeras authorization of assigned analysis sets is required for the final analysis. Agreement and authorization of 
participants included/excluded from each analysis set will be obtained prior to the unblinding activities of the 
study. 
In view of the final analysis, the following analysis sets will be defined: 
5.1. RANDOMIZED ANALYSIS SET 
The Randomized analysis set is defined as all participants who were randomized to either H4:IC31, Placebo or 
BCG regardless of whether they received a study vaccination. 
A participant will be programmatically included in the Randomized analysis set if the participant has a 
randomization number in the clinical study database, as recorded on the System Enrollment eCRF. 
For analyses and presentations based on Randomized analysis set, participants will be analyzed according to the 
treatment assigned regardless of the treatment received. 
5.2. SAFETY ANALYSIS SET 
The Safety analysis set is defined as all participants included in the Randomized analysis set who received at 
least one dose of study vaccination (H4:IC31, Placebo or BCG). 


 
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A participant will be included in the Safety analysis set if the participant: 
• 
Is included in the Randomized analysis set. 
AND 
• 
Has at least one date of study vaccination in the database as recorded on the Exposure (Day 0) or 
Exposure (Day 56) eCRF. 
For analyses and presentations based on the Safety analysis set, participants will be analyzed according to the 
treatment received regardless of the treatment allocated (that is ‘as treated’): 
• 
For all presentations, including by dose at onset presentations, participants who received a different 
treatment to that allocated, but the same consistent treatment for all relevant doses, analyze and 
present participants according to the treatment received (both overall and per dose at onset). 
• 
Participants who received a different treatment to that allocated, but not the same consistent treatment 
for all relevant doses (mixed treatment regimen), will be analyzed as described below: 
o 
For all presentations, with the exception of by dose at onset presentations, analyze and present 
participants according to the following vaccine hierarchy: H4:IC31 followed by BCG and then 
placebo. 
o 
For all by dose at onset presentations, analyze participants according to the treatment received for 
each dose at onset; therefore, participants are analyzed and presented according to different 
treatments per the relevant doses.  
The Safety analysis set will be used as the primary analysis set for all safety and immunogenicity analyses. 
5.3. MODIFIED INTENT-TO-TREAT ANALYSIS SET 
The Modified Intent-to-treat analysis set is defined as all participants included in the Safety analysis set who 
had a negative QFT-GIT test result at Day 84. 
A participant will be programmatically included in the Modified Intent-to-treat analysis set if the participant: 
• 
Is included in the Safety analysis set. 
AND 
• 
Has a negative QFT-GIT test result at the Day 84 visit. 
For analyses and presentations based on the Modified Intent-to-treat analysis set, participants will be analyzed 
according to the treatment assigned regardless of the treatment received. 
The Modified Intent-to-treat analysis set will be used for efficacy analyses. 


 
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5.4. ADJUSTED MODIFIED INTENT-TO-TREAT (FIRST RECALL POSITIVE OMIT 
ANALYSIS SET) 
This analysis set will omit all participants whose first positive QFT-GIT result came from a recalled tube (refer to 
Section 3.3: Changes to Analysis from Protocol). This analysis set will therefore include the same participants 
who are included within the mITT set except for the following nine participants, all of whom had their first 
positive QFT-GIT result from a recalled tube: 
044002-00288 
044002-01409 
044002-00433 
044002-01438 
044002-00587 
044002-01506 
044002-00883 
044002-02063 
044002-00923 
Some of the primary efficacy analyses will also be performed using the Adjusted Modified Intent-to-Treat 
analysis set as a sensitivity analysis. 
5.5. PER PROTOCOL ANALYSIS SET 
The Per Protocol analysis set is defined as all participants in the Modified Intent-to-treat analysis set who 
received all relevant doses of BCG, H4: IC31 or Placebo and had no major protocol deviations. 
A participant will be included in the Per Protocol analysis set if the participant: 
• 
Is included in the Modified Intent-to-treat analysis set. 
• 
Received all relevant doses, based on the date of study vaccination as recorded on Exposure (Day 0) and 
Exposure (Day 56) eCRFs: 
o 
BCG: Day 0. 
o 
H4:IC31 or Placebo: Day 0 and Day 56. 
• 
Did not have any major protocol deviations as recorded on the Protocol Deviation eCRF and agreed with 
Aeras prior to the relevant unblinding activities of the analysis/study. 
• 
Did not have a treatment deviation as identified in collaboration with the clinical trial manager (CTM) in 
accordance with the Aeras/QuintilesIMS Process Instruction Treatment Deviation Identification and 
Authorization document. 
• 
Out-of-window QFT-GIT positive results should not be included in the analysis for the PP analysis set. 
Refer to the relevant Blind Data Review (BDR) documentation for additional details. 


 
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The Per Protocol analysis set will be based on the participant level flag assigned in Appendix 1 of the BDR 
report, as authorized by Aeras prior to DB lock. 
Efficacy analyses will also be performed using the Per Protocol analysis set as a sensitivity analysis. 
 
 


 
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6. 
GENERAL CONSIDERATIONS 
6.1. REFERENCE START DATE AND STUDY DAY 
Reference start date is defined as the date of first study vaccination and equals Day 0. Study day will be 
calculated relative to the reference start date and will be used to show start or stop day of an assessment or 
event. Study day will be calculated as follows: 
• 
If the date of the assessment/event is prior to, on or after the reference start date: 
o 
Study day = (date of assessment/event – reference start date). 
Unless otherwise specified, if the date of the assessment/event is partial or missing, study day and any 
corresponding durations will not be calculated and will be presented as missing in the listings. 
Some presentations include the study day relative to the most recent vaccination (presented as relative day) 
and will be calculated as follows: 
• 
If the date of the assessment/event is on or after the most recent vaccination: 
o 
Relative day = (date of assessment/event – date of most recent vaccination). 
6.2. BASELINE 
Unless otherwise specified, Baseline is defined as the last non-missing assessment (scheduled or unscheduled) 
prior to the first study vaccination. In the case where the last non-missing assessment and the reference start 
date coincide, that assessment will be considered pre-vaccination (Baseline). For example, if vital signs, 
laboratory or electrocardiogram assessments fall on the date of first study vaccination and the time of the 
assessment is missing, the applicable assessment will be considered as Baseline. However, adverse events (AEs) 
starting on the reference start date (date of first study vaccination) will be considered as treatment-emergent, 
therefore post-baseline. 
6.3. RETESTS, UNSCHEDULED VISITS AND PREMATURE DISCONTINUATION 
DATA 
In general, for by-visit summaries, data recorded at the scheduled visit will be presented. Unscheduled 
assessments will not be included in by-visit summaries, but may contribute to the Baseline value. 
In the case of a retest (same visit number assigned), the latest available test result as provided in the data 
transfer for that visit/time point will be used for by-visit summaries. 
Listings will include scheduled, unscheduled and premature discontinuation data. No mapping of premature 
discontinuation data will be performed. 
 
 


 
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6.4. STATISTICAL TESTS 
Unless otherwise specified: 
• 
The default summary statistics for quantitative variables will be as follows: 
o 
Number of participants in each category (n). 
o 
Mean. 
o 
Standard deviation (SD). 
o 
Median. 
o 
Minimum. 
o 
Maximum. 
• 
The default summary statistics for qualitative variables will be as follows: 
o 
Number of participants in each category (n). 
o 
Percentage of participants in each category (%), presented relative to either one of the following: 
▪ 
Total number of participants in the relevant analysis set. 
▪ 
Total number of participants in the relevant analysis set, with assessments available (observed 
cases). 
• 
In the event of missing assessments, a ‘Missing’ category showing the number of participants with 
missing assessments at each level of summarization will be presented. 
• 
Methods for determining confidence intervals (CIs) will be specified in each relevant section.   
6.5. COMMON CALCULATIONS 
The following common calculations are defined: 
• 
Change from Baseline = (Result at relevant post-baseline time point – Result at Baseline). 
• 
Most recent vaccination: 
o 
Calculate the difference in the start date of the assessment/event and all available study vaccination 
dates: 
▪ 
Days since dose x: (AE start date – study vaccination dose x date [where x = 1, 2 {not applicable 
for BCG}])  
o 
For the previous calculation, only consider those doses where the days since dose x is ≥ 0 (therefore 
excluding the doses occurring after the relevant assessment/event) and determine the dose (1, 2 
[not applicable for BCG]) for which the days since dose x (as calculated in the previous step) is a 
minimum. 
o 
Assign the dose selected in the previous step as the most recent vaccination as the specific dose 
determined in the previous calculation. 
• 
Change from pre-vaccination: 
o 
Result at Study Day x – results at relevant pre-vaccination time point. 
o 
If a result/value is missing, change from pre-vaccination result will be presented as missing in the 
listing. That is, no imputation for a missing change will be performed.  


 
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• 
Shift in toxicity grade from pre-vaccination: 
o 
Toxicity grade at Study Day x – toxicity grade at relevant pre-vaccination time point. 
o 
If a toxicity grade is missing, shift in toxicity grade from pre-vaccination will be presented as missing 
in the listing. That is, no imputation for a missing shift will be performed.  
o 
If no toxicity grade is assigned to a specific visit the grade will be regarded as grade 0 for calculation 
purposes. 
6.6. SOFTWARE VERSION 
All analyses will be conducted using SAS® Version 9.4. 
7. 
STATISTICAL CONSIDERATIONS 
7.1. ADJUSTMENTS FOR COVARIATES AND FACTORS TO BE INCLUDED IN 
ANALYSES 
Not applicable. 
7.2. MULTICENTER STUDIES 
This study is being conducted by multiple Investigators at multiple schools/regions in South Africa. No 
adjustment for school/region effect will be made. Generally, all schools/regions will be pooled by cohort and 
treatment group for analysis purposes. 
7.3. MISSING DATA 
Missing data will not be imputed. For the handling of partial or missing AE start dates, refer to APPENDIX 2: 
Partial Date Conventions. 
7.4. MULTIPLE COMPARISONS/MULTIPLICITY 
There will be no adjustment for multiplicity to control the Type 1 error rate over the two tests associated with 
the primary efficacy endpoint. The rationale for not performing this adjustment is related to the fact that the 
tests pertain to efficacy assessments of two unrelated vaccines (H4:IC31 and BCG). If these two evaluations 
were performed in two separate trials then no adjustment for multiplicity would be made. Thus, one should 
not be compelled to adjust for this multiplicity simply because the two evaluations are performed 
administratively in a single trial. 


 
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In addition, this is considered an intermediate test of biological effect study in the development of the vaccine 
and no definitive claims are being sought. 
Multiplicity adjustment for immunology data will be performed by means of the Holm (Step-Down Bonferroni) 
method.  
7.5. EXAMINATION OF SUBGROUPS 
No subgroup analyses will be performed. 
8. 
OUTPUT PRESENTATIONS 
APPENDIX 1: Programming Conventions for Outputs describes the conventions for presentation of data in the 
TLFs. 
The TLF shells provided together with this SAP describe the presentations for the final analysis and therefore 
the format and content of the TLFs to be provided by QuintilesIMS Biostatistics. 
Note that verbatim terms, specifications (for example the reason a specific assessment was not done) and all 
variables in the TLF shells that contain the suffix (eCRF) contain verbatim text that may include spelling 
mistakes. Verbatim text will be presented in the listings ‘as is’ and no manual ‘hard-coding’ corrections of such 
data will be made. 
For table presentations, the following treatment groups and cohorts will be presented: 
• 
Treatment Group: 
o 
Placebo. 
o 
H4:IC31. 
o 
BCG. 
o 
Total. 
• 
Cohort (if applicable): 
o 
All participants. 
o 
Safety and Immunogenicity Cohort. 
o 
Correlates Cohort. 
9. 
DISPOSITION AND PREMATURE DISCONTINUATION 
All participants who provide informed consent and were randomized to a treatment group and cohort will be 
accounted for in this study. 
Participant disposition will be presented for the Randomized analysis set (unless otherwise specified). 
The following tables are planned for presentation: 
• 
Randomization by site.  
• 
Participant disposition and primary reason for premature study discontinuation. 


 
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• 
Participant disposition and primary reason for premature treatment discontinuation (Safety analysis set). 
• 
Protocol deviations. 
• 
Analysis sets. 
The following listings are planned for presentation: 
• 
Participant disposition.  
• 
Protocol deviations.  
• 
Inclusion and/or exclusion exceptions at Study Day 0. 
• 
Analysis sets. 
The following variable will be collected at Screening on the System Enrollment eCRF: 
• 
InForm participant number (12-digit): 
o 
First three digits are the standard study indicator (‘044’). 
o 
Next four digits are the study site indicator followed by a dash (‘002-‘ or ‘014-‘). 
o 
Last five digits are a sequential participant number (00001 to 03000), uniquely assigned per 
participant. 
The following disposition variables, as presented in the planned TLF shells, will be collected at Screening on the 
Informed Consent/Cohort Assignment eCRF: 
• 
Date of informed consent. 
• 
Date of informed assent. 
• 
Date of randomization. 
The following disposition variables, as presented in the planned TLF shells, will be collected during the study on 
the Disposition Study eCRF: 
• 
Whether the participant completed the study: 
o 
Yes. 
o 
No. 
• 
Primary reason for premature study discontinuation: 
o 
Lost to follow-up. 
o 
Withdrew consent. 
o 
Death. 
o 
Adverse Event. 
o 
Protocol deviation. 
o 
Study terminated by sponsor. 
o 
Other (specify). 
• 
Date of study completion/premature discontinuation. 
The following disposition variables, as presented in the planned TLF shells, will be collected during the study on 
the Disposition Treatment eCRF: 
• 
Whether the participant completed study treatment: 
o 
Yes. 
o 
No. 


 
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• 
Primary reason for premature study treatment discontinuation: 
o 
Lost to follow-up. 
o 
Withdrew consent. 
o 
Death. 
o 
Adverse Event (specify). 
o 
Protocol deviation. 
o 
Study terminated by sponsor. 
o 
Physician decision (specify). 
o 
Other (specify). 
• 
Date of treatment completion/premature discontinuation.  
The following variable will be collected on the Exposure eCRF: 
• 
Date of study vaccination(s). 
The following protocol deviation variables, as presented in the planned TLF shells, will be collected throughout 
the study on the Protocol Deviations eCRF: 
• 
Visit to which the protocol deviation is relevant. 
• 
Protocol deviation category: 
o 
Informed consent not properly obtained. 
o 
Did not meet inclusion criteria or met exclusion criteria but entered in the study. 
o 
Developed withdrawal criteria during the study but not withdrawn. 
o 
Study vaccine dosing deviation. 
o 
Procedure(s) performed out of window.  
o 
Missed procedure(s). 
o 
Unintended unblinding (not applicable to the BCG treatment group). 
o 
Serious adverse event (SAE)/immediately reportable event (IRE) reporting. 
o 
Other (specify). 
• 
Date of protocol deviation. 
• 
Protocol deviation description. 
The following variables, as presented in the planned TLF shells, will be collected prior to Screening on the 
Inclusion/Exclusion Criteria (IE) eCRF and collected at Day 0 on the Eligibility Verification (IEV) eCRF: 
• 
Category: 
o 
Inclusion.  
o 
Exclusion.  
• 
Criterion number.  
• 
Date of criterion exception noted.  
The following study visit variable, as presented in the planned TLF shells, will be collected throughout the study 
on the Date of Visit eCRF: 
• 
Date of visit. 


 
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The following variables, as presented in the planned TLF shells, will be imported from an Aeras approved EXCEL 
spreadsheet created for the review of major protocol deviations as well as inclusion of participants in each 
analysis set: 
• 
For each analysis set, included in analysis set (refer to Section 5: Analysis Sets for relevant analysis sets 
and definitions): 
o 
Yes. 
o 
No. 
• 
For each analysis set, reason(s) for exclusion from analysis set (refer to Section 5: Analysis Sets for 
possible reasons for exclusion from each analysis set). 
9.1. DERIVATIONS 
Based on the aforementioned, the following variables, as presented in the planned TLF shells, will be derived: 
• 
Treatment unblinded: 
o 
Based on unintended unblinding on the Protocol Deviation eCRF.  
• 
Update the InForm participant number to the participant ID (8-digit): 
o 
Replace four digits from digit 4 to 7 with the 3-digit study site number, by removing the first digit 
(‘002-’ → ’02-’ and ‘014-’ → ’14-’). 
o 
Last five digits kept as is. 
This participant ID will be used in all the listings, as presented in the relevant TLF shells. 
• 
Total number of doses:  
o 
Per participant, determine the number of doses received.  
o 
If date of vaccination is available, the vaccination will be considered as administered.  
• 
Age (years), sex, race: 
o 
Refer to Section 10: Demographic and Other Baseline Characteristics and Section 10.1: Derivations 
for details. 
10. DEMOGRAPHIC AND OTHER BASELINE CHARACTERISTICS 
No inferential statistical testing will be performed for demographic or other Baseline characteristics. 
The following table is planned for presentation (based on the Safety analysis set): 
• 
Demographics and other Baselines characteristics. 
o 
Presented by means of the default summary statistics for quantitative and qualitative variables. 
o 
Percentage (%) of participants will be calculated relative to the total number of participants in the 
Safety analysis set per treatment group with data available (observed case). 
o 
This table will be repeated for the Modified Intent-to-treat analysis set, Per Protocol analysis set as 
well as the Safety and Immunogenicity Cohort (Safety analysis set). 
The following listing is planned for presentation (based on the Randomized analysis set): 
• 
Demographics and other Baseline characteristics.  


 
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The following demographic variables, as presented in the planned TLF shells, will be collected at Screening on 
the Demography eCRF: 
• 
Date of birth. 
• 
Sex: 
o 
Male (M). 
o 
Female (F). 
• 
Race: 
o 
Asian. 
o 
Black. 
o 
White. 
o 
Coloured. 
o 
Other (specify). 
The following variable, as presented in the planned TLF shells, will be collected at Screening on the Informed 
Consent/Cohort Assignment eCRF: 
• 
Cohort assignment: 
o 
Safety and Immunogenicity Cohort. 
o 
Correlates Cohort. 
The following demographic variable, as presented in the planned TLF shells, will be collected at Screening on 
the School Selection eCRF: 
• 
School: 
o 
Drostdy. 
o 
Esselenpark Sekondêr. 
o 
Hexvallei Hoërskool. 
o 
Hexvallei Sekondêr. 
o 
Hoërskool Breërivier. 
o 
HS Goudini. 
o 
Montana Hoërskool. 
o 
Van Cutsem. 
o 
Vusisizwe. 
o 
Worcester Gimnasium. 
o 
Worcester Sekondêr. 
o 
Other (specify). 
The following demographic variable, as presented in the planned TLF shells, will be collected at Screening on 
the Region Selection eCRF: 
• 
Region: 
o 
Athlone. 
o 
Gugulethu. 
o 
New Crossroads. 
o 
Nyanga. 
o 
Old Crossroads. 
o 
Philippi East. 
o 
Philippi West. 
o 
Other (specify). 


 
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The following Baseline characteristic, as presented in the planned TLF shells, will be collected on the Height and 
Weight eCRF: 
• 
Baseline body mass index (BMI) (kg/m2). 
10.1. DERIVATIONS 
Based on the aforementioned the following variable, as presented in the planned TLF shells, will be derived: 
• 
Age (years): 
o 
Calculated relative to the reference start date (Day 0 vaccination). 
o 
Age (years) = (date of Day 0 vaccination) – (date of birth)/365.25. 
All listings, as presented in the relevant TLF shells, will include the following demographic information: 
• 
Age (years). 
• 
Sex. 
• 
Race. 
These demographic variables will be concatenated to form one demographic variable in the following format: 
• 
Age (years)/Sex/Race: 
o 
For example: 17/M/WHITE. 
11. MEDICAL HISTORY  
Medical History will be presented for the Randomized analysis set. 
The following listing is planned for presentation:  
• 
Medical history.  
Medical history will be coded using Medical Dictionary for Regulatory Activities (MedDRA) Version 16.1. 
The following medical history variables, as presented in the planned TLF shells, will be collected on the Medical 
History (MH) and Interval Medical History (IMH) eCRFs during the study:  
• 
Verbatim term (eCRF).  
• 
Date started.  
• 
Date stopped or ongoing.  
• 
Comment (eCRF).  
The following medical history coding variables, as presented in the planned TLF shells, will be included in the 
data transfers received from Data Management: 
• 
System Organ Class.  
• 
Preferred Term.  


 
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No imputation of partial or missing dates will be performed for medical history and study days will not be 
presented for these cases.  
12. PRIOR AND CONCOMITANT MEDICATIONS 
Concomitant medications will be presented for the Randomized analysis set. 
The following listing is planned for presentation:  
• 
Concomitant medications.  
Concomitant medication will be coded with World Health Organization – Drug Dictionary (WHO-DD) Version 
01DEC2013. 
The following concomitant medication variables, as presented in the planned TLF shells, will be collected on the 
Prior and Concomitant Medication (CM) eCRF: 
• 
Verbatim term (eCRF). 
• 
Date started.  
• 
Date stopped or ongoing.  
• 
Dose and unit per administration.  
• 
Frequency.  
• 
Route of administration.  
• 
Indication.  
The following medication coding variables, as presented in the planned TLF shells, will be included in the data 
transfers received from Data Management: 
• 
Preferred Term.  
• 
Anatomical Therapeutic Chemical (ATC) Level 1 term.  
• 
ATC Level 3 term.  
No imputation of partial or missing dates will be performed for medications and study days will not be 
presented for these cases.  
13. STUDY VACCINATION 
Administration of the study vaccine(s) will be presented for the Safety analysis set. 
The following table is planned for presentation: 
• 
Study vaccination administration: 
o 
Presented by treatment group by means of the default summary statistics for qualitative variables. 
o 
Percentage (%) of participants will be calculated relative to the total number of participants in the 
Safety analysis set per treatment group. 


 
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o 
The number and percentage (%) of participants who received all scheduled vaccinations will be 
presented. 
o 
The number and percentage (%) of participants who received the second vaccination within the 
protocol-defined window will be presented. The denominator for the % will be based on the total 
number of participants in the Safety analysis set per treatment group (excluding BCG).  
The following listing is planned for presentation (based on the Randomized analysis set): 
• 
Study vaccination administration.  
Study vaccinations will be administered on the study days as specified for each treatment group in Table 1: 
Planned Enrollment by Treatment Group and Cohort. 
The following exposure variables, as presented in the planned TLF shells, will be collected on the Exposure Day 
0 and Exposure Day 56 eCRFs: 
• 
Study vaccination administered: 
o 
Yes. 
o 
No. 
• 
Date and time of study vaccination. 
• 
Full volume administered: 
o 
Yes. 
o 
No. 
• 
Location (arm): 
o 
Left. 
o 
Right. 
The following treatment deviation variables, as presented in the planned TLF shells, will be collected in 
collaboration with the clinical trial manager (CTM) with the Aeras/Quintiles Process Instruction: Treatment 
Deviation Identification and Authorization document: 
• 
In accordance with randomization: 
o 
Yes. 
o 
No. 
• 
Out of study window: 
o 
Yes. 
o 
No. 
13.1. DERIVATIONS 
Based on the aforementioned the following variables, as presented in the planned TLF shells, will be derived: 
• 
Participants who received all scheduled vaccinations: 
o 
Yes: 
▪ 
For BCG treatment group participants if Day 0 study vaccination date is available. 
▪ 
For H4:IC31 or Placebo treatment group participants if Day 0 and Day 56 study vaccination dates 
are both available. 


 
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o 
No: 
▪ 
For BCG treatment group participants if Day 0 study vaccination date is not available. 
▪ 
For H4:IC31 or Placebo treatment group participants if Day 0 and/or Day 56 vaccination dates are 
not available. 
• 
Study day of the Day 56 vaccination: 
o 
Calculated relative to Day 0 vaccination (refer to Section 6.1: Reference Start Date and Study Day). 
• 
Participants who received the second vaccination within the protocol-defined window: 
o 
This is only applicable to the H4:IC31 and Placebo participants. 
o 
The allowable window for the Day 56 visit is 56-2 days until 56+14 days from the Day 0 vaccination. 
o 
Yes: 
▪ 
Study day of the Day 56 vaccination >= 54. 
AND 
▪ 
Study day of the Day 56 vaccination <= 70. 
o 
No: 
▪ 
Study day of the Day 56 vaccination < 54. 
OR 
▪ 
Study day of the Day 56 vaccination > 70. 
14. EFFICACY OUTCOMES 
14.1. PRIMARY EFFICACY 
14.1.1. 
PRIMARY EFFICACY ENDPOINT 
The primary efficacy endpoint is defined as QuantiFERON TB Gold-in-tube (QFT-GIT) conversion, at any time 
point after Day 84 through end of follow-up, from negative at Screening to positive, using the manufacturer’s 
recommended threshold of ≥ 0.35 IU/mL. 
14.1.2. 
ANALYSIS OF PRIMARY EFFICACY ENDPOINT 
The following tables will be presented for the Modified Intent-to-treat analysis set: 
• 
QFT-GIT results at each visit: 
o 
Number and percentage (%) of participants in each category per visit: 
▪ 
Negative.  
▪ 
Positive.  
▪ 
Missing.  
o 
Percentage (%) of participants will be calculated relative to the total number of participants in the 
Modified Intent-to-treat analysis set who have QFT-GIT assessments available per treatment group 
per visit.  


 
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• 
Compliance to QFT-GIT testing: 
o 
Number and percentage (%) of participants in each compliance category per visit: 
▪ 
Yes. 
▪ 
No. 
▪ 
Missing. 
o 
Percentage (%) of participants will be calculated relative to the total number of participants in the 
Modified Intent-to-treat analysis set expected to have a QFT-GIT assessment available per treatment 
group per visit (N1). 
• 
QFT-GIT conversion rates, primary endpoint (therefore endpoints occurring after Day 84): 
o 
Number and percentage (%) of participants in each conversion status category: 
▪ 
Yes. 
▪ 
No. 
▪ 
Missing. 
o 
Hazard ratio (H4:IC31/comparator) presented: 
▪ 
Point estimates (based on the Cox regression model). 
▪ 
P-values for the comparison of the estimated hazard ratios of QFT-GIT conversion based on a 
one-sided log-rank test. 
• 
H4:IC31 versus Placebo. 
• 
BCG versus Placebo. 
• 
H4:IC31 versus BCG (at the request of the sponsor’s partner). 
o 
Vaccine efficacy presented: 
▪ 
Point estimate (based on the hazard ratio estimates). 
▪ 
80% two-sided CI based on the hazard ratio estimate from the Cox regression model. 
o 
This table will be repeated for the Per Protocol, Adjusted Modified Intent-to-treat (First Recall 
Positive Omit) analysis sets. This table will also be repeated for the adjudication approach (refer to 
Section 3.3: Changes to Analysis from Protocol). 
o 
As a sensitivity analysis the content of this table will be repeated in a separate table using interval 
censoring for the MITT analysis set.  
• 
First QFT-GIT conversion by visit: 
o 
Number and percentage (%) of participants with first QFT-GIT conversion at each of the scheduled 
visits. 
o 
Number of participants at risk for first conversion at each scheduled visit. 
o 
At risk defined as participants with: 
▪ 
A negative QFT-GIT result at the preceding visit without a previous positive QFT-GIT result. 
▪ 
A QFT-GIT result available at the relevant visit. 
o 
This table will be repeated for the Per Protocol analysis set. 
• 
Follow-up time until first QFT-GIT conversion: 
o 
Number of participants contributing to the total follow-up time (at risk). 
o 
Cumulative number of participants: 
▪ 
Censored (not due to a primary endpoint at the time of the last available non-positive QFT-GIT 
result). 
▪ 
With an event (primary endpoint). 
o 
The total follow-up time (person-months), as measured from Randomization (1 month = 4 weeks). 
o 
The Kaplan-Meier event rate presented at each relevant post-Day 84 visit. 
o 
This table will be repeated for the Per Protocol analysis set. 


 
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• 
Primary endpoints by school/region: 
o 
Number and percentage (%) of participants who experienced a primary endpoint in each 
school/region. 
• 
Primary endpoints by site and sex: 
o 
Number and percentage (%) of participants who experienced a primary endpoint in each site/sex. 
The following table will be presented for the Randomized analysis set: 
• 
QFT-GIT conversion rates including all post-Day 0 conversions: 
o 
Number and percentage (%) of participants in each conversion status category: 
▪ 
Yes. 
▪ 
No. 
▪ 
Missing. 
o 
Two groups of conversion time points included: 
▪ 
Any QFT-GIT conversion (early [Day 84] or post-Day 84) (at risk participants include all 
participants). 
▪ 
Early (Day 84) conversion (at risk participants include all participants). 
o 
Hazard ratio (H4:IC31/comparator) presented: 
▪ 
Point estimates (based on the Cox regression model). 
▪ 
P-values for the comparison of the estimated hazard ratios of QFT-GIT conversion based on a 
one-sided log-rank test. 
• 
H4:IC31 versus Placebo. 
• 
BCG versus Placebo. 
• 
H4:IC31 versus BCG (included at the request of the sponsor’s partner). 
o 
Vaccine efficacy presented: 
▪ 
Point estimate (based on the hazard ratio estimates). 
▪ 
80% two-sided CI based on the hazard ratio estimate from the Cox regression model. 
The following figures will be presented for the Modified Intent-to-treat analysis set: 
• 
Time on study: 
o 
Kaplan-Meier plot per treatment group. 
o 
Cumulative follow-up time as measured from randomization (1 month = 4 weeks). 
o 
Event is defined as whichever occurs first: 
▪ 
Initial QFT-GIT conversion. 
▪ 
Early termination/study completion. 
o 
This figure will be repeated for the Per Protocol analysis set. 
• 
Time to first post-Day 84 QFT-GIT conversion, primary endpoint: 
o 
Kaplan-Meier plots per treatment group for the following: 
▪ 
Time to first post-Day 84 QFT-GIT conversion. 
o 
Time to first QFT-GIT conversion will be presented in months, measured from Day 84 (1 month = 4 
weeks). 
▪ 
Censored (at the time of the last available non-positive QFT-GIT result). 
o 
This figure will be repeated for the Per Protocol analysis set. 
• 
Time to first post-Day 84 QFT-GIT conversion (interval-censored sensitivity analysis): 
o 
Survival function estimate plot per treatment group based on a life-table analysis. 
o 
Time to QFT-GIT conversion measured from Day 84. 
o 
Time to conversion will be presented in 6-month intervals. 


 
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The following figure will be presented for the Randomized analysis set: 
• 
Time to First QFT-GIT Conversion (Early or Post-Day 84): 
o 
Kaplan-Meier plots per treatment group for the following: 
▪ 
Time to any QFT-GIT conversion (early or post-Day 84) measured from Day 0. 
o 
Time to QFT-GIT conversion will be presented in months, measured from Randomization (1 month = 
4 weeks). 
▪ 
Censored (not due to a primary endpoint at the time of the last available non-positive QFT-GIT 
result). 
The following listing will be presented for the Randomized analysis set: 
• 
QuantiFERON results. 
The following QuantiFERON variables, as presented in the planned TLF shells, will be collected on the 
QuantiFERON and QuantiFERON Re-valuation eCRFs: 
• 
Date of sample collection. 
• 
Qualitative results: 
o 
Negative. 
o 
Positive. 
• 
Quantitative Result: 
o 
QFT-Nil (IU/mL). 
o 
QFT-Ag (IU/mL). 
o 
QFT-Mitogen (IU/mL).  
14.1.3. 
DERIVATIONS 
Based on the aforementioned, the following variables, as presented in the planned TLF shells, will be derived 
for post-vaccination visits and/or time points: 
• 
QFT-GIT testing compliance derived per visit: 
o 
Yes: QFT-GIT assessment date is within the protocol-defined visit window. 
o 
No: QFT-GIT assessment date is outside the protocol-defined visit window. 
o 
Missing: The participant is expected to have a QFT-GIT assessment available. 
o 
Refer to APPENDIX 8: Protocol-defined Visit Windows for the relevant protocol-defined visit 
windows. 
• 
QFT-GIT result classification at each visit (use the retest result if available). The Investigator’s assessment 
will be used, which is based on the algorithm below.   
QFT-GIT Result Classification  
QFTNil  
QFTAg - QFTNil 
QFTMitogen - QFTNil 
Positive 
≤ 8.0 
 
≥ 0.35 (primary 
analysis threshold) 
and ≥ 25% of QFTNil 
value 
Any  
Negative 
< 0.35 (primary 
≥0.5 


 
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QFT-GIT Result Classification  
QFTNil  
QFTAg - QFTNil 
QFTMitogen - QFTNil 
Indeterminate 
analysis threshold) OR 
≥ 0.35 (primary 
analysis threshold) 
and < 25% of QFTNil 
value 
< 0.5 
Indeterminate 
> 8.0 
Any 
Any 
• 
If the QFT-GIT result is indeterminate at any of the visits Day 84, M6, M9, M12, M15, M18, M21, M24, 
M27 or M30, the QFT-GIT test will be repeated (QUANT2 form in the eCRF used to record the result). If 
the repeat result is negative or indeterminate, the participant will continue with the study visits as if the 
participant did not convert, and the result for the corresponding visit will be deemed “negative”. 
• 
Any QFT-GIT conversion status: 
o 
The QFT-GIT conversion status is assigned hierarchically in the order provided below: 
▪ 
Yes: Participant has at least one positive QFT-GIT result classification, regardless of missing or 
indeterminate results at any other visit and regardless of at which visit the positive QFT-GIT 
result is recorded.  
▪ 
Post-Day 84 QFT-GIT conversion: First positive QFT-GIT result classification recorded post-
Day 84. 
▪ 
Early QFT-GIT conversion: First positive QFT-GIT result classification recorded at Day 84. 
▪ 
No: Participant has no positive QFT-GIT result classification and the last available QFT-GIT 
result classification is negative. 
▪ 
Missing: Participant has no positive/negative/indeterminate QFT-GIT result classification 
available. 
• 
Censored: 
o 
Participants who do not experience a primary endpoint or early conversion (early conversion is only 
applicable to the analyses based on the Randomized analysis set), will be censored at the date of 
their last available, non-positive QFT-GIT result. 
o 
The following SAS code will be implemented for all Kaplan-Meier analyses, where: 
PROC LIFETEST DATA = <>; 
 
TIME Months*Censored(1); 
 
TEST Treatment_Group; 
RUN; 
▪ 
Censored = 1 represents censored participant. 
▪ 
Censored = 0 represents participant who experienced an event. 
▪ 
Months represent the available follow-up time (months) per participant since: 
• 
Day 84 (applicable to analyses based on the mITT and PP analysis set). 
• 
Randomization (applicable to analyses based on the Randomized analysis set). 
• 
Event: 
o 
Participants who experience a primary endpoint or early conversion will be indicated as having an 
event (depending on the relevant table, early converters will be included/excluded from the 
participants experiencing an event). 


 
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• 
Available follow-up time (months) until first QFT-GIT conversion: 
o 
The number of months of follow-up time will be derived for each participant based on the date of 
randomization/Day 84 (depending on the relevant table) and either the censor or event date. 
o 
1 month = 4 weeks. 
o 
For the sensitivity analysis the follow-up time will be rounded to the nearest 6-month interval for 
any of the following visits: 
▪ Month 6. 
▪ Month 12 
▪ Month 18. 
▪ Month 24. 
o 
For the sensitivity analysis the following SAS code will be implemented: 
PROC LIFETEST DATA = <>  METHOD=lt INTERVALS=(0 to 2 by 0.5)  
PLOTS=(s); 
 
 
TIME Years*Censored(1); 
 
 
FREQ count; 
RUN; 
• 
Vaccine efficacy is derived as: 
o 
𝑉𝐸 =  1 –  𝐻𝑎𝑧𝑎𝑟𝑑 𝑅𝑎𝑡𝑒 𝑅𝑎𝑡𝑖𝑜 (𝑉𝑎𝑐𝑐𝑖𝑛𝑒/𝑐𝑜𝑚𝑝𝑎𝑟𝑎𝑡𝑜𝑟) 
o 
The hazard ratio is estimated based on the following Cox regression model: 
PROC PHREG DATA = <>; 
 
 
MODEL Time to Event (Months) * Censor (1) = Treatment Group; 
RUN; 
▪ Censor = 1 represents censored participant. 
▪ Censor = 0 represents participant who experienced an event. 
▪ Months represent the available follow-up time (months) per participant since randomization. 
o 
The 80% CI is based on the two-sided hazard ratio confidence interval and will be derived as: 
▪ 𝑉Elower limit  =  1 −Hazard ratioupper limit 
▪ VEupper limit = 1 −𝐻𝑎𝑧𝑎𝑟𝑑 𝑟𝑎𝑡𝑖𝑜𝑙𝑜𝑤𝑒𝑟 𝑙𝑖𝑚𝑖𝑡 
o 
The 95% CI is based on the two-sided hazard ratio confidence interval.  
• 
For the sensitivity analysis, interval censoring will be used based on the following code: 
PROC ICPHREG DATA = <>; 
MODEL (left, right) = Treatment Group /b=pch(NINTERVAL=X); 
RUN; 
▪ Left represents left censoring value in months.  
▪ Right represents right censoring value in months. 
▪ Left and right censoring values are dependent on where the event is identified: 
 
Visit at Which Primary Event 
Identified 
Left Censoring Value 
Right Censoring Value 
M6 
3 
6 
M12 
6 
12 
M18 
12 
18 
M24 
18 
24 
 


 
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14.2. SECONDARY EFFICACY 
14.2.1. 
SECONDARY EFFICACY ENDPOINT 
The secondary efficacy endpoint is defined as sustained QFT-GIT conversion without reversion: 
• 
Occurrence of a primary endpoint. 
• 
Sustained conversion without a change in QFT-GIT from positive to negative through 6 months after QFT-
GIT conversion, also using the threshold of 0.35 IU/mL. 
• 
Sustained conversion without a change in QFT-GIT from positive to negative through 6 months after QFT-
GIT conversion, using the alternative threshold of 0.2 at any time point prior to conversion and > 0.70 at 
initial post-Day 84 QFT-GIT conversion, and maintains QFT-GIT > 0.7 for 6 months after initial conversion. 
A participant will be considered as having a secondary endpoint (at the first conversion date), denoted here as 
‘6-month sustained conversion (per protocol)’, if that participant had three consecutive positive QFT-GIT 
results after Day 84. 
14.2.2. 
ANALYSIS OF SECONDARY EFFICACY ENDPOINT 
The following table will be presented for the Modified Intent-to-treat analysis set: 
• 
QFT-GIT 6-month sustained conversion rates: 
o 
Number and percentage (%) of participants in each conversion status category: 
▪ 
Yes. 
▪ 
No. 
▪ 
Missing. 
o 
Hazard ratio (H4:IC31/comparator) presented: 
▪ 
Point estimates (based on the Cox regression model). 
▪ 
P-values for the comparison of the estimated hazard ratios of QFT-GIT 6-month sustained 
conversion based on a one-sided log-rank test. 
• 
H4:IC31 versus Placebo. 
• 
BCG versus Placebo. 
• 
H4:IC31 versus BCG (at the request of the sponsor’s partner). 
o 
Vaccine efficacy presented: 
▪ 
Point estimate (based on the hazard ratio estimate). 
▪ 
80% two-sided CI based on the hazard ratio estimate from the Cox regression model. 
o 
This table will be repeated for the alternative threshold value.  
o 
These tables (based on the original threshold as well as the alternative threshold) will be repeated 
for the Per Protocol analysis set. This table (based on the original threshold) will also be repeated for 
the Adjusted Modified Intent-to-treat (First Recall Positive Omit) analysis set and the adjudication 
approach (refer to Section 3.3: Changes to Analysis from Protocol). 
o 
As a sensitivity analysis both of these tables (original threshold and the alternative threshold 
outputs) will be repeated in separate tables using interval censoring for the MITT analysis set.  


 
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The following figures will be presented for the Modified Intent-to-treat analysis set: 
• 
Time to 6-month sustained QuantiFERON conversion: 
o 
Kaplan-Meier plots per treatment group for the following: 
▪ 
Time to first of three consecutive post-Day 84 QFT-GIT positive results. 
o 
Time to sustained QFT-GIT conversion will be presented in months, measured from Day 84 (1 month 
= 4 weeks) to the date of the first of three consecutively positive QFT-GIT results. 
o 
Participants who do not convert are censored at their last negative QFT-GIT.  
o 
This figure will be repeated for the alternative threshold value.  
o 
Both these figures (original and alternative threshold outputs) will be repeated for the Per Protocol 
analysis set. 
The following QuantiFERON variables, as presented in the planned TLF shells, will be collected on the 
QuantiFERON and QuantiFERON Re-valuation eCRFs: 
• 
Date of sample collection. 
• 
Qualitative results: 
o 
Negative. 
o 
Positive. 
• 
Quantitative Result: 
o 
QFT-Nil (IU/mL). 
o 
QFT-Ag (IU/mL). 
o 
QFT-Mitogen (IU/mL). 
14.2.3. 
DERIVATIONS 
Based on the aforementioned, the following variables (refer to Section 14.1.2: Analysis of Primary Efficacy 
Endpoint), as presented in the planned TLF shells, will be derived at 3 and 6 months post first QFT-GIT 
conversion for participants who have a post-Day 84 QFT-GIT conversion (primary endpoint): 
• 
QFT-GIT result classification (refer to Section 14.1.3: Derivations). 
• 
QFT-GIT 6-month sustained conversion status: 
o 
Yes: At both 3 and 6 months following first QFT-GIT conversion, QFT-GIT result classifications are 
derived as positive. 
o 
No: At 3 and/or 6 months following first QFT-GIT conversion, QFT-GIT result classification is derived 
as negative. 
o 
Missing: None of the aforementioned QFT-GIT result classifications (yes, no) has been assigned at 3 
or 6 months post-conversion, QFT-GIT result classification is derived as missing. 
• 
Censored: 
o 
Participants who do not experience an initial QFT-GIT conversion will be censored at the date of their 
last available, non-positive QFT result. 
o 
Converters who do not have a sustained conversion will be censored at their last available QFT-GIT 
result.  
o 
Converters with only a 3-month post-conversion result and no 6-month post-conversion result will 
be censored at: 
▪ 
The initial conversion visit if the 3-month post-conversion result is positive.  


 
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▪ 
The 3-month post-conversion visit if that 3-month post-conversion result is negative. 
o 
The following SAS code will be implemented for all Kaplan-Meier analyses, where: 
PROC LIFETEST DATA = <>; 
 
TIME Months*Censored(1); 
 
TEST Treatment_Group; 
RUN; 
▪ 
Censored = 1 represents censored participants. 
▪ 
Censored = 0 represented participants who experienced an event (sustained QFT-GIT 
conversion). 
▪ 
Months represent the available follow-up time (months) per participant since Day 84. 
• 
Event: 
o 
Participants, who experience a sustained QFT-GIT conversion for three consecutive visits will be 
indicated as having an event. 
• 
Vaccine efficacy is derived as: 
o 
𝑉𝐸 =  1 –  𝐻𝑎𝑧𝑎𝑟𝑑 𝑅𝑎𝑡𝑖𝑜 (𝑉𝑎𝑐𝑐𝑖𝑛𝑒/𝑐𝑜𝑚𝑝𝑎𝑟𝑎𝑡𝑜𝑟) 
o 
The hazard ratio is estimated based on the following Cox regression model: 
PROC PHREG DATA = <>; 
 
 
MODEL Time to Event (Months) * Censor (1) = Treatment Group; 
RUN; 
▪ 
Censor = 1 represents censored participant. 
▪ 
Censor = 0 represents participant who experienced an event. 
▪ 
Months represent the available follow-up time (months) per participant since Day 84. 
o 
The 80% CI is based on the two-sided hazard ratio confidence interval and will be derived as: 
▪ 
𝑉Elower limit  =  1 −Hazard ratioupper limit 
▪ 
VEupper limit = 1 −𝐻𝑎𝑧𝑎𝑟𝑑 𝑟𝑎𝑡𝑖𝑜𝑙𝑜𝑤𝑒𝑟 𝑙𝑖𝑚𝑖𝑡 
o 
The 95% CI is based on the two-sided hazard ratio confidence interval.  
14.3. EXPLORATORY EFFICACY 
14.3.1. 
EXPLORATORY EFFICACY ENDPOINT 
The following exploratory analyses will be performed: 
• 
Prevention of Mtb infection as measured by rates of reversion.  
• 
Effect of alternative QFT-GIT test threshold values on rates of QFT-GIT conversion and reversion.  
• 
To explore trends in QFT-GIT prolonged/sustained conversions and late reversions (i.e., more than 6 
months post initial conversion) in early QFT-GIT converters (i.e., among those who converted at Month 6 
or Month 12 of follow-up). 
Some analyses are based on subsets of participants determined by a post-randomization event, and some 
involve participants who experience differential follow-up. Given these types of analyses have the potential to 
be biased, additional supportive analyses, such as principal stratification, may be utilized to better understand 
a potential vaccine effect.  


 
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14.3.2. 
ANALYSIS OF EXPLORATORY EFFICACY ENDPOINT 
The following tables will be presented for the Modified Intent-to-treat analysis set: 
• 
QuantiFERON levels (IU/mL) upon and following first positive QFT-GIT result: 
o 
Default summary statistics for QFT-GIT results at the following time points: 
▪ 
First positive QFT-GIT result: 
• 
The p-value based on the Mann-Whitney-Wilcoxon test will be used for pair-wise 
comparisons between treatment groups. 
▪ 
3 months-post first positive QFT-GIT result: 
• 
Number and percentage (%) of participants with a negative QFT-GIT. 
▪ 
6 months-post first positive QFT-GIT result: 
• 
Number and percentage (%) of participants with a negative QFT-GIT. 
o 
This table is to include all participants with at least one positive post-Day 84 QFT-GIT result. 
o 
This table will be repeated for the Per Protocol analysis set. 
• 
First QuantiFERON reversion: 
o 
Number of participants with first QFT-GIT negative value within the following categories will be 
included: 
▪ 
Within 3 months post-conversion. 
▪ 
Within 6 months post-conversion. 
▪ 
At any time during follow-up.  
o 
At risk defined as participants with: 
▪ 
A primary endpoint. 
▪ 
A QFT-GIT result available at the relevant visit within 6 months (for the first two lines) or at any 
time post-initial conversion (for the third line) after the primary endpoint occurrence. 
o 
This table will be repeated for the Per Protocol analysis set. 
• 
Alternative QFT-GIT thresholds: 
o 
Number and percentage of participants who EVER experienced the following alternative threshold 
values (i.e., at any post-Day 84 timepoint): 
▪ 
≥ 0.20. 
▪ 
< 0.2 at any time point prior to conversion and > 0.70 at any time point post-Day 84. 
▪ 
> 0.70. 
▪ 
> 4.0.  
o 
Vaccine efficacy presented: 
▪ 
Point estimate (based on the conditional binomial distribution).  
▪ 
95% CI (based on the conditional binomial Clopper-Pearson method with mid-p correction).    
o 
This table will be repeated for the Per Protocol analysis set.  
• 
Trends in QFT-GIT sustained conversions and end-of-study reversions: 
o 
Number and percentage (%) of participants in each category: 
▪ 
Participants who were negative at Month 24 or the Month 24 call-back visit. Percentages (%) will 
be calculated based on the number of participants with non-missing Month 24 or Month 24 call-
back values.  
▪ 
Participants who never converted or converted with reversion. Percentages (%) will be calculated 
based on the number of participants who did not have two missing QFT-GIT values within 6 
months of post-initial conversion.  


 
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▪ 
Participants who never converted or converted with reversion and have non-positive (i.e., 
negative or missing) six-month post-initial-conversion QFT-GIT values. Percentages (%) will be 
calculated based on the number of participants who did not have two missing QFT-GIT values 
within 6 months of post-initial conversion.  
▪ 
Participants who never converted or converted with reversion and have non-positive (i.e., 
negative or missing) end of study QFT-GIT values. Percentages (%) will be calculated based on the 
number of participants who did not have the following combination of QFT-GIT values: +/m/m, 
+/+/m/m or +/m/m/m where + = positive and m = missing.  
o 
Comparisons of proportions between treatment groups for each category will be performed based on 
the Pearson chi-square test.  
o 
Vaccine efficacy presented: 
▪ 
Point estimate (based on the conditional binomial Clopper-Pearson method with mid-p 
correction).  
▪ 
95% CI (based on the conditional binomial Clopper-Pearson method with mid-p correction).    
• 
QFT-GIT end of study sustained conversion rates: 
o 
Number and percentage (%) of participants in each conversion status category: 
▪ 
Yes. 
▪ 
No. 
▪ 
Missing. 
o 
Hazard ratio (H4:IC31/comparator) presented: 
▪ 
Point estimates (based on the Cox regression model). 
▪ 
P-values for the comparison of the estimated hazard ratios of QFT-GIT end of study sustained 
conversion based on a one-sided log-rank test. 
• 
H4:IC31 versus Placebo. 
• 
BCG versus Placebo. 
• 
H4:IC31 versus BCG (at the request of the sponsor’s partner). 
o 
Vaccine efficacy presented: 
▪ 
Point estimate (based on the hazard ratio estimate). 
▪ 
80% two-sided CI based on the hazard ratio estimate from the Cox regression model. 
o 
This table will be repeated for the Per Protocol analysis set. 
• 
Participants who converted, reverted, and time to conversion among reverters and non-reverters: 
o 
Number and percentage (%) of participants who: 
▪ 
Converted.  
▪ 
Converted and later reverted.  
▪ 
Converted and did not revert.  
o 
The median time and the quartile 1 (Q1) and 3 (Q3) values will be displayed for: 
▪ 
Time to conversion among all converters.  
▪ 
Time to conversion among all reverters.  
▪ 
Time to conversion among non-reverters.  
o 
This table will be repeated for the Per Protocol analysis set.  
• 
QFT-GIT positivity patterns over time from initial conversion timepoint onwards: 
o 
Number and percentage (%) of participants, summarized for the following patterns in the order 
specified below: 
▪ 
+/+/+/+ 
▪ 
+/+/+ 
▪ 
+/+/m 


 
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▪ 
+/m/+ 
▪ 
+/m/m/m 
▪ 
+/+/+/- 
▪ 
+/-/+/+ 
▪ 
+/+/-/+ 
▪ 
+/+/- 
▪ 
+/-/+ 
▪ 
+/-/m 
▪ 
+/-/-/+ 
▪ 
+/-/- 
▪ 
+/-/+/- 
▪ 
+/m/-/- 
▪ 
+/-/-/- 
▪ 
-/-/-/- 
▪ 
This table will also be repeated for the Per Protocol analysis set. 
• 
Clinical TB diagnosis: 
o 
Number and percentage of participants in each of the following categories: 
▪ 
Participants who received at least one TB work-up: 
▪ 
Bacteriologically-confirmed TB. 
▪ 
Clinical TB (based on the Investigator’s assessment). 
o 
The total number of TB work-ups performed. 
o 
This table will be repeated for the Per Protocol analysis set. 
The following figures will be presented for the Modified Intent-to-treat analysis set: 
• 
Quantitative QFT-GIT results following initial conversion: 
o 
Box-and-whisker plots per treatment group for the following: 
▪ 
QFT-GIT result at 0, 3 and 6 months since first QFT-GIT conversion (reference line at 0.35 IU/mL 
to be included). 
▪ 
QFT-GIT Change from Baseline at 3 and 6 months since first QFT-GIT conversion (reference line 
at 0 IU/mL to be included). 
▪ 
The number of participants at 3 and 6 months since post-initial conversion whose results fell 
below 0.35 should also be displayed.  
o 
This figure will include all participants with at least one positive QFT-GIT result post-Day 84. 
o 
This figure will be repeated for the Per Protocol analysis set. 
• 
Time to first QuantiFERON reversion: 
o 
Kaplan-Meier plot per treatment group. 
o 
Time to first reversion will be presented in months, measured from primary endpoint occurrence (1 
month = 4 weeks). 
o 
Participants who do not revert (Month 24 call-back visit also considered) will be censored at the time 
of the last positive QFT-GIT result. 
o 
This figure will be repeated for the Per Protocol analysis set. 
• 
Time to first QFT-GIT conversion among those who ever reverted (Month 24 call-back visit also 
considered): 
o 
Kaplan-Meier plots per treatment group for the following: 
▪ 
Time to post-Day 84 QFT-GIT conversion. 


 
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o 
Time to QFT-GIT conversion will be presented in months, measured from Day 84 (1 month = 4 
weeks) to the date of the first QFT-GIT conversion of those who ever reverted. 
o 
Participants who do not convert are censored at their last negative QFT-GIT.  
This figure will be repeated for the Per Protocol analysis set. 
• 
Time to end of study sustained QuantiFERON conversion: 
o 
Kaplan-Meier plots per treatment group for the following: 
▪ 
Time to first of three (M18 or M24 initial conversion) or four (M6 or M12 initial conversion) 
consecutive post-Day 84 QFT-GIT positive results. 
o 
Time to sustained QFT-GIT conversion will be presented in months, measured from Day 84 (1 month 
= 4 weeks) to the date of the first of three (M18 or M24 initial conversion) or four (M6 or M12 initial 
conversion) consecutively positive QFT-GIT results. 
o 
Participants who do not convert are censored at their last non-positive QFT-GIT.  
o 
This figure will be repeated for the Per Protocol analysis set. 
The following listing will be presented for the randomized analysis set: 
• 
Clinical TB diagnosis and assessment.  
• 
QuantiFERON positivity patterns over time.  
• 
Participants who QFT-GIT converted, reverted and then converted again.  
The following QuantiFERON variables, as presented in the planned TLF shells, will be collected on the 
QuantiFERON and QuantiFERON Re-valuation eCRFs: 
• 
Date of sample collection. 
• 
Qualitative results: 
o 
Negative. 
o 
Positive. 
• 
Quantitative Result: 
o 
QFT-Nil (IU/mL). 
o 
QFT-Ag (IU/mL). 
o 
QFT-Mitogen (IU/mL). 
The following TB diagnosis variables, as presented in the planned TLF shells, will be collected on the TB 
Assessments – First Sputum Sample and TB Assessments – Second Sputum Sample eCRFs: 
• 
Date of sputum sample collection. 
• 
Sputum smear result: 
o 
Acid- Fast Bacilli (AFB) seen: 
▪ 
Scanty. 
▪ 
1+. 
▪ 
2+. 
▪ 
3+. 
o 
No AFB seen. 
o 
No result. 
• 
Sputum culture result: 
o 
Mycobacterium positive. 
o 
Mycobacterium negative. 


 
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o 
Contaminated. 
o 
No result. 
• 
Differential test for positive Culture: 
o 
MTB. 
o 
Mycobacterium other than tuberculosis (MOTT) (specify). 
• 
GeneXpert result: 
o 
Invalid. 
o 
No result. 
o 
MTB not detected. 
o 
MTB detected. 
The following TB diagnosis variables, as presented in the planned TLF shells, will be collected on the TB Clinical 
Evidence eCRF: 
• 
TB clinical signs and symptoms: 
o 
Unexplained cough for more than two weeks. 
o 
Fever for more than two weeks. 
o 
Night sweats for more than two weeks. 
o 
Unexplained weight loss. 
o 
Recent history of Hemoptysis. 
o 
Other (specify). 
The following TB diagnosis variables, as presented in the planned TLF shells, will be collected on the Chest X-
Ray eCRF: 
• 
Chest x-ray performed: 
o 
Yes. 
o 
No. 
• 
Chest x-ray result: 
o 
Normal. 
o 
Abnormal. 
The following clinical TB diagnosis variable, as presented in the planned TLF shells, will be collected on the TB 
Case Assessment eCRF: 
o 
Was a clinical case of TB diagnosed? 
14.3.3. 
DERIVATIONS 
Based on the aforementioned, the following variables, as presented in the planned TLF shells, will be derived 
for post-vaccination visits and/or time points: 
• 
QFT-GIT result classification (refer to Section 14.1.3: Derivations). Substitute the primary analysis 
threshold value with the alternative threshold value.  
• 
Available follow-up time (months) until QFT-GIT: 
o 
Months represent the available follow-up time (months) per participant since first QFT-GIT 
conversion. 


 
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• 
Negative at Month 24: 
o 
Participants who are negative at Month 24 or who initially converted at Month 6 or Month 12 and 
are negative at the ≥ 24 Month call-back visit.  
• 
Prevention of conversion OR conversion with reversion: 
o 
Participants who are consistently negative OR 
o 
Participants who have a negative QFT-GIT value within six months after their initial conversion (i.e., 
+/-/-, +/+/-, +/-/+, +/m/-, +/-/m).  
• 
Prevention of conversion OR conversion with reversion and non-positive (i.e.  negative or missing) QFT-
GIT results at six-months post-initial-conversion: 
o 
Participants who are consistently negative OR  
o 
Participants who have a negative QFT-GIT value at the last available testing timepoint within six 
months after their initial conversion (i.e., +/-/-, +/+/-, +/m/-, +/-/m). 
• 
Prevention of conversion OR conversion with reversion and non-positive (i.e., negative or missing) 
QFT-GIT results at the end of the study: 
o 
Participants who are consistently  negative throughout the study OR 
o 
Participants who initially converted at Month 18 or Month 24 (i.e., +/-/-, +/+/-, +/m/-, +/-/m) and 
later reverted OR 
o 
Participants who initially converted at Month 6 or Month 12 and have a call-back Month 24 visit that 
is negative (i.e., +/-/-/-, +/+/-/-, +/+/+/-, +/-/+/-, +/-/m/-, +/m/-/-, +/m/m/-, +/+/m/-).  
• 
QFT-GIT end of study sustained conversion status: 
o 
Yes: At 3 and 6 months following first QFT-GIT conversion AND at the Month 24 call-back visit (if 
applicable), QFT-GIT result classifications are derived as positive. 
o 
No: At 3 and/or 6 months following first QFT-GIT conversion and/or at the Month 24 call-back visit (if 
applicable), QFT-GIT result classification is derived as negative. 
o 
Missing: None of the aforementioned QFT-GIT result classifications (yes, no) has been assigned at 3 
or 6 months post-conversion or at the Month 24 call-back visit, QFT-GIT result classification is 
derived as missing. 
• 
The confidence interval for VE is calculated using the conditional binomial method (Clopper-Pearson 
method with mid-p correction) as follows: 
The number of participants with a conversion (nA) in one of the treatment groups (for example H4:IC31) 
and the number (nB) of participants with a conversion in another treatment group (for example Placebo), 
are assumed to have Independent Poisson distributions with rate parameters λA and λB, respectively, but 
inference is conditional on the total number of conversions, n = nA + nB. Conditional on n, nA has a 
Binomial distribution, Bin(n,π), where 
π = 
λA FA 
= 
θ FA 
λA FA + λB FB 
θ FA + FB 
 
 θ = λA\λB and FA and FB are the total lengths of follow-up across all subjects in the relevant treatment 
groups (Treatment Group A/Treatment Group B), that is,  F = Σ (Duration of VE Follow-up Period 
[Days]). 


 
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A point estimate and exact two-sides 95% CI (πL,πU) for π is then calculated (Clopper - Pearson method 
with mid-p correction) based on the number of participants with a conversion in one of the treatment 
groups (for example H4:IC31), relative  to the total number of participants n = nA + nB who had at a 
conversion in either of the two compared treatment groups.  The following SAS® code will be used: 
 
PROC FREQ DATA = <>; 
 
 
TABLE Events / BINOMIAL (midp P=0.05); 
RUN; 
Based on the exact 95% CI for π, an exact 95% CI (θL,θU), for the rate ratio θ is then calculated by solving 
the equations: 
θL FA 
= 
πL 
θL FA + FB 
 
θU FA 
= 
πU 
θU FA + FB 
 
Solving θL and θU yields: 
θL = ( 
πL 
)( 
FB ) 
1 - πL 
FA 
 
 
 
θU = ( 
πU 
)( 
FB ) 
1 - πU 
FA 
 
Finally, the exact 95% CI (VEL, VEU) for VE is calculated as 
VEL = (1 – θU)  
VEU = (1 – θL)  
In summary, the VE point estimate and confidence limits for the aforementioned binomial proportion (π) 
will be substituted in the following formula in order to obtain the corresponding confidence limits for the 
VE: 
VE  = ( 
(π) FB 
)  
(1 – π)FA 
 


 
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Where the lower confidence limit for VE is obtained by substituting the upper confidence limit for π in 
the above formulae, and the upper confidence limit for VE is obtained by substituting the lower 
confidence limit for π in the above formula.  
• 
Bacteriologically confirmed TB: 
o 
Positive culture result (Mycobacterium tuberculosis positive) 
OR 
o 
Positive GeneXpert result (MTB detected) (with or without clinical symptoms and/or positive chest x-
ray). 
 
 


 
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15. IMMUNOGENICITY 
15.1. PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMC) INTRACELLULAR 
CYTOKINE STAINING (ICS) 
The following tables are planned for presentation (based on the Safety analysis set, in the Safety and 
Immunogenicity Cohort): 
• 
Percent antigen-specific T cell negative control-subtracted ANY cytokine response and change from 
Baseline. 
o 
Only the derived ANY cytokine combination will be included in this table. 
o 
This table will include descriptive statistics and 95% confidence interval (CI) for the median based on 
order statistics at each applicable visit as well as the change from Baseline per T cell and stimulation 
antigen (including a ‘total’ stimulation antigen for Ag85B and TB10.4 combined). 
o 
This table will be repeated for the negative-control ANY cytokine combination.  
• 
Percent antigen-specific T cell negative-control subtracted Positive (G,2) cytokine response and change 
from Baseline.  
o 
Only the derived Positive (G,2) cytokine combination will be included in this table. 
o 
This table will include descriptive statistics and 95% confidence interval (CI) for the median based on 
order statistics at each applicable visit as well as the change from Baseline per T cell and stimulation 
antigen (including a ‘total’ stimulation antigen). 
o 
This table will be repeated for the negative control Positive (G,2) cytokine combination. 
• 
Number (percentage) of responders based on the derived Positive (G,2) cytokine combination. 
o 
This is to include the number and percentage of responders (Refer to Section 15.1.2: Responder 
Status) at each visit per T cell and stimulation antigen. 
o 
Percentages are based on the number of participants in the Safety analysis set within each treatment 
group with data available to determine responder status.  
o 
P-values are also to be included and are based on a Fisher’s EXACT test comparing each of the 
treatment groups.  
• 
Comparisons of median negative control-subtracted ANY cytokine response. 
o 
Only the derived ANY cytokine combination will be included in this table. 
o 
This table will include the number of participants who have a negative control-subtracted negative-
set-to-zero value available as well as the median of these values.  
o 
P-values based on the Wilcoxon-Mann Whitney test for pair-wise comparisons between treatment 
groups will be performed.  
• 
Comparisons of median negative control-subtracted Positive (G,2) cytokine response. 
o 
Only the derived Positive (G,2) cytokine combination will be included in this table. 
o 
This table will include the number of participants who have a negative control-subtracted negative-
set-to-zero value available as well as the median of these values.  
o 
P-values based on the Wilcoxon-Mann Whitney test for pair-wise comparisons between treatment 
groups will be performed.  


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 52 of 91 
The following figures are planned for presentation (based on the Modified Intent-to-treat analysis set): 
• 
Antigen-specific negative control-subtracted ANY cytokine response: 
o 
Dot plot of antigen-specific negative control-subtracted ANY cytokine response per study day. 
o 
One figure will be prepared for each stimulation antigen, including the derived total (Ag85B and 
TB10.4) stimulation antigen response; the median response will be represented by a horizontal line 
per treatment group and study day. 
o 
Separate plots will be created per T cell (CD4+/CD8+). 
o 
This figure will be repeated for the negative control ANY cytokine combination. 
o 
Different symbols will be used to differentiate between responders (closed circles), non-responders 
(cross) and missing responder status (triangle) as based on the derived Positive (G,2) cytokine 
combination responder status.  
• 
Antigen-specific negative control-subtracted Positive (G,2) cytokine response: 
o 
Dot plot of antigen-specific negative control-subtracted Positive (G,2) cytokine response per study 
day. 
o 
One figure will be prepared for each stimulation antigen, including the derived total (Ag85B and 
TB10.4) stimulation antigen response; the median response will be represented by a horizontal line 
per treatment group and study day. 
o 
Separate plots will be created per T cell (CD4+/CD8+). 
o 
This figure will be repeated for the negative control Positive (G,2) cytokine combination.  
o 
Different symbols will be used to differentiate between responders (closed circles), non-responders 
(cross) and missing responder status (triangle) as based on the derived Positive (G,2) cytokine 
combination responder status.  
• 
Antigen-specific negative control-subtracted ANY cytokine median response: 
o 
Line plot of antigen-specific negative control-subtracted ANY cytokine median response per study 
day. Bars will be included, representing the first and third quartile per study day. 
o 
One figure will be prepared for each stimulation antigen, including the derived total (Ag85B and 
TB10.4) stimulation antigen response; the median response will be represented by a dot and 
connected with a horizontal line across study days per treatment group. 
o 
Separate plots will be created per T cell (CD4+/CD8+). 
o 
This figure will be repeated for the negative control ANY cytokine combination. 
• 
Antigen-specific negative control-subtracted Positive (G,2) cytokine median response: 
o 
Line plot of antigen-specific negative control-subtracted Positive (G,2) cytokine median response per 
study day. Bars will be included, representing the first and third quartile per study day. 
o 
One figure will be prepared for each stimulation antigen, including the derived total (Ag85B and 
TB10.4) stimulation antigen response; the median response will be represented by a dot and 
connected with a horizontal line across study days per treatment group. 
o 
Separate plots will be created per T cell (CD4+/CD8+). 
o 
This figure will be repeated for the negative control Positive (G,2) cytokine combination. 
• 
Total (Ag85B and TB10.4) negative control-subtracted ANY cytokine response: 
o 
Stacked bar plot of each stimulation antigen’s median negative control-subtracted ANY cytokine 
response per study day. 
o 
Separate plots will be created per T cell (CD4+/CD8+). 
• 
Total (Ag85B and TB10.4) negative control-subtracted Positive (G,2) cytokine response: 


 
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o 
Stacked bar plot of each stimulation antigen’s median negative control-subtracted Positive (G,2) 
cytokine response per study day. 
o 
Separate plots will be created per T cell (CD4+/CD8+). 
• 
Antigen-specific negative control-subtracted polyfunctional profile plots: 
o 
Separate plots will be created per cytokine combinations of interest: 
▪ 
CD4+: IFN-g, IL-2, TNF, CD154. 
▪ 
CD4+: IFN-g, IL-2, IL-17, IL-22. 
▪ 
CD8+: IFN-g, IL-2, TNF, CD107a. 
o 
Dot plot of participants’ negative control-subtracted cytokine response per single-gate on the left 
hand side. Dot plot of participants’ negative control-subtracted ANY cytokine response on the right 
hand side. The median response will be represented by a horizontal line per treatment group and 
cytokine combination. 
o 
These figures will be repeated for the negative control. 
The following listings are planned for presentation (based on the Safety analysis set): 
• 
Percent antigen-specific T cell negative control-subtracted cytokine response and change from Baseline: 
o 
Only derived cytokine combinations will be included in this listing. 
• 
Percent antigen-specific T cell negative control-subtracted ANY cytokine response and total (Ag85B and 
TB10.4) vaccine response: 
o 
Only derived cytokine combinations will be included in this listing. 
• 
ICS responder status based on the Positive (G,2) response: 
o 
The responder status for each antigen will be displayed, including the overall status.  
The following PBMC ICS variables, as presented in the planned TLF shells, will be contained in the data received 
from Aeras: 
• 
Stimulation antigen: 
o 
Ag85B. 
o 
BCG. 
o 
TB10.4. 
o 
Negative control: 
▪ 
DMSO for Ag85B and TB10.4. 
▪ 
R10 for BCG.  
• 
T cell: 
o 
CD4+. 
o 
CD8+. 
• 
Cell counts and percentages (including a total cell count) for each possible combination (alone and in 
combination with all other cytokines) of the following cytokines per stimulation antigen, study day and 
participant: 
o 
IFN-𝛾 (+/-). 
o 
TNF (+/-). 
o 
IL-2 (+/-). 
o 
IL-17A (+/-). 
o 
IL-22 (+/-). 
o 
CD154 (+/-). 
o 
CD107a (+/-). 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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15.1.1. 
DERIVATIONS 
15.1.1.1. 
Negative control-subtracted Cytokine Response 
The negative control results are sometimes referred to as the background data. For all immunology 
presentations, the negative control results are subtracted from the active stimulation antigen results. This 
result is referred to as the negative control-subtracted response. 
15.1.1.2. 
Single Positive CD107a 
The single positive CD107a cytokine combination (for example: APBNGN2N4N17NTN or 
APBNGN2N17N22NTN), per antigen, is excluded entirely from the Analysis Data Model (ADaM) ICS Result 
(ADZI) dataset.  
Since there are 126* cytokine combinations (128* cytokine combinations in total, excluding the all negative 
and the single positive CD107a cytokine combinations) available to be presented on a polyfunctional figure, it 
would not be possible to present all combinations on a single figure. Therefore, a subset of four cytokines is to 
be presented on one figure. The figure presenting the IFN-g, IL-2, TNF and CD107a subset only include 14 
polyfunctional combinations on the x-axis instead of the usual 15 combinations, since the last column (IFN-g - 
IL-2- TNF- CD107a+) is omitted from the figure.  
15.1.1.3. 
Total Number of Cells 
The ICS Total number of cells per participant, per day, per T cell, per antigen does include all 128* cytokine 
combinations, therefore the single positive CD107a cytokine combination is included in the ICS Total number of 
cells. The ICS Total number of cells for each cell type is available in the RAW data file and is not recalculated as 
the sum of all cytokine combinations. The ICS Total number of cells from the RAW dataset:  
• 
Time/Singlets/Viable CD3+/Lymphocytes/Aggregate 1/Aggregate 2/CD4+,Count for CD4.  
• 
Time/Singlets/Viable CD3+/Lymphocytes/Aggregate 1/Aggregate 2/CD8+,Count for CD8.  
15.1.1.4. 
ANY Response 
The ‘ANY’ (%) result per participant, T cell type, stimulation antigen and visit are derived as: 
• 
∑𝐶𝑒𝑙𝑙 𝑝𝑒𝑟𝑐𝑒𝑛𝑡𝑎𝑔𝑒𝑠 (%) 𝑡ℎ𝑎𝑡 𝑟𝑒𝑠𝑝𝑜𝑛𝑑 (𝑝𝑜𝑠𝑖𝑡𝑖𝑣𝑒) 𝑎𝑡 𝑡ℎ𝑒 𝑐𝑦𝑡𝑜𝑘𝑖𝑛𝑒 𝑙𝑒𝑣𝑒𝑙. 
The single positive CD107a and all negative combinations should be excluded from this derivation. 
15.1.1.5. 
Total Stimulation Antigen Result 
The total stimulation antigen result per participant, T cell type and visit is derived as: 
• 
∑′𝐴𝑁𝑌′(%) 𝑟𝑒𝑠𝑢𝑙𝑡𝑠 𝑜𝑓 𝐴𝑔85𝐵 𝑎𝑛𝑑 𝑇𝐵10.4  𝑎𝑛𝑡𝑖𝑔𝑒𝑛𝑠 (𝑒𝑥𝑐𝑙𝑢𝑑𝑖𝑛𝑔 𝑛𝑒𝑔𝑎𝑡𝑖𝑣𝑒 𝑐𝑜𝑛𝑡𝑟𝑜𝑙 𝑎𝑛𝑑 𝐵𝐶𝐺). 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 55 of 91 
15.1.1.6. 
Positive (G,2) 
The ‘Positive (G,2)’ (%) result per participant, T cell type, stimulation antigen and visit is derived as: 
• 
∑𝐶𝑒𝑙𝑙 𝑝𝑒𝑟𝑐𝑒𝑛𝑡𝑎𝑔𝑒𝑠 (%) 𝑤ℎ𝑒𝑟𝑒 𝐼𝐹𝑁−𝑔 𝑎𝑛𝑑/𝑜𝑟 𝐼𝐿−2 𝑎𝑟𝑒 𝑝𝑜𝑠𝑖𝑡𝑖𝑣𝑒. 
15.1.2. 
RESPONDER STATUS 
To determine the responder status the following steps will be followed: 
• 
Per participant, T cell type, stimulation antigen (including negative control) and visit, derive: 
o 
Positive (G,2). 
o 
Negative (G,2): 
▪ 
Total Cell Count – Positive (G,2). 
o 
The applicable cytokines are: IFN-g (G) and IL-2 (2).  
• 
These results and the negative control results (positive and negative cell counts) will then be compared 
at each visit per participant, T cell type and stimulation antigen, using the Fisher’s EXACT test (refer to 
Section 5.8.5: Fisher’s Exact Test in the Figure guidelines).  
• 
P-values will be adjusted for multiplicity by using PROC MULTTEST (Holm Method). The p-values are 
adjusted across antigens, therefore at a T cell type, by visit-level for each participant. 
• 
Step 1: Determine whether the number of cells responding to the active antigen stimulation is 
statistically significantly more than the number of cells responding to the negative control stimulation at 
Baseline (Day 0).  
o 
One-sided adjusted p-value >= 0.05 indicates that the number of cells responding to the active 
antigen stimulation is not statistically more than the number of cells responding to the negative 
control stimulation.  
▪ 
Participant is classified as a non-responder at Baseline (Day 0).  
o 
One-sided adjusted p-value < 0.05 indicates that the number of cells responding to the active 
antigen stimulation is statistically more than the number of cells responding to the negative control 
stimulation.  
▪ 
Participant is classified as a responder at Baseline (Day 0).  
• 
Step 2: Determine whether the number of cells responding to the active antigen stimulation is 
statistically significantly more than the number of cells responding to the negative control stimulation, at 
Day 70.  
o 
One-sided adjusted p-value >= 0.05 indicates that the number of cells responding to the active 
antigen stimulation is not statistically more than the number of cells responding to the negative 
control stimulation.  
▪ 
Participant is classified as a non-responder at Day 70.  
o 
One-sided adjusted p-value < 0.05 indicates that the number of cells responding to the active 
antigen stimulation is statistically more than the number of cells responding to the negative control 
stimulation  
▪ 
Participant is classified as a responder at Day 70.  
 
 


 
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• 
Determine the responder status post-Baseline when taking Baseline (Day 0) into account:  
Baseline (Determined in Step 
1) 
Day 70 (Determined in Step 
2) 
Responder Status 
Non-responder 
Non-responder 
Non-responder 
Non-responder 
Responder 
Responder 
Responder 
Non-responder 
Non-responder 
Responder 
Responder 
Determine responder status by 
means of the odds ratio  
• 
Determine the odds ratio for results at Baseline. If the negative control positive count is zero, the odds 
ratio will be undefined. In these cases the positive negative control count should be imputed to 1 in 
order to determine an odds ratio.  
• 
Determine the odds ratio for results at Day 70. If the negative control count is zero, the odds ratio will be 
undefined. In these cases the positive negative control count should be imputed to 1 in order to 
determine an odds ratio.  
• 
Perform the Breslow-Day test: 
o 
Test for homogeneity of odds ratio at Day 70 and Baseline (Day 0).  
o 
P-value >= 0.05 indicates that the odds ratio at Baseline (Day 0) and Day 70 do not differ statistically 
significantly.  
▪ 
Participant classified as a non-responder at the relevant post-Baseline visit.  
o 
P-value < 0.05 indicates that the odds ratio at Baseline (Day 0) and Day 70 visit differ statistically 
significantly.  
▪ 
Difference in log odds ratio should be evaluated in order to determine the responder status 
at Day 70.  
• 
Calculate the difference in the log of the odds ratios at Day 70 and Baseline (Day 0).  
o 
Difference in log odds ratio = log(odds ratio at Day 70) - log(odds ratio at Baseline [Day 0]).  
• 
Determine the responder status based on the Breslow-Day and difference in log odds ratio for cases 
where the Baseline (Day 0) and Day 70 visit was determined as responders: 
Breslow-Day test p-value 
Difference in log odds ratio 
Responder status 
>= 0.05 
<0 
Non-responder 
>=0.05 
>0 
Non-responder 
<0.05 
<0 
Non-responder 
<0.05 
>0 
Responder 
The aforementioned steps will be implemented to determine the data record-level response per 
participant, T cell type, stimulation antigen (excluding negative control) and visit. In order to determine 
the response per participant at a day-level and therefore across T cell type and stimulation antigen, the 
following conventions will be used: 
• 
Per participant and visit, if all records have a record-level responder status of non-responder, the day 
level responder status is determined as a non-responder.  


 
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• 
Per participant and visit, if at least one of the records has a record-level responder status of responder, 
the day-level responder status is determined as responder.  
The overall responder status is determined as a single responder status per participant across T cell types, 
stimulation antigens and visits and the following conventions will be used to determine the overall responder 
status: 
• 
Per participants, if all days have a day-level responder status of non-responder the overall responder 
status is determined as non-responder.  
• 
Per participants, if at least one of the days has a day-level responder status of responder, the overall 
responder status is determined as responder.  
As the Baseline responder status is used to determine the post-Baseline responder status, it might be possible 
that not all participants have sufficient data at Baseline (Day 0) or the relevant post-Baseline visit (Day 70) to 
determine the responder status. These participants will be presented in the figures by means of a different 
symbol (i.e. triangle) to indicate that the responder status could not be determined.  
15.2. WHOLE BLOOD ICS 
The following tables are planned for presentation (based on the Safety analysis set, in the Safety and 
Immunogenicity Cohort). Aeras will be responsible for performing this analysis.  
• 
Percent antigen-specific T cell negative control-subtracted cytokine response and change from Baseline. 
• 
Number (percentage) of responders.  
• 
Comparisons of median negative control-subtracted cytokine response. 
These tables will be provided for the following cytokine combinations (including repeat tables for the negative 
control for each cytokine combination): 
• 
CD4 cells producing Positive (G,2,T,17,22) cytokine combination. 
• 
CD4 cells producing Positive (G,2,T) cytokine combination. 
• 
CD8 cells producing Positive (G,2,T,17,22) cytokine combination. 
• 
CD8 cells producing Positive (G,2,T) cytokine combination.  
• 
CD4 cells producing Positive (17) cytokine combination.  
• 
CD4 cells producing Positive (22) cytokine combination.  
The following listings are planned for presentation (based on the Safety analysis set): 
• 
Percent antigen-specific T cell negative control-subtracted cytokine response and change from Baseline: 
o 
Only derived cytokine combinations will be included in this listing. 
• 
Percent antigen-specific T cell negative control-subtracted cytokine response and total (Ag85B and 
TB10.4) vaccine response: 
o 
Only derived cytokine combinations will be included in this listing. 
• 
ICS responder status based on each derived cytokine response: 
o 
The responder status for each antigen will be displayed, including the overall status.  


 
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The following whole blood ICS variables, as presented in the planned TLF shells, will be contained in the data: 
• 
Stimulation antigen: 
o 
Ag85B. 
o 
BCG. 
o 
TB10.4. 
o 
Negative control:  
▪ 
UNS. 
• 
T cell: 
o 
CD4+. 
o 
CD8+. 
• 
Cell counts and percentages (including a total cell count) of the following cytokines per stimulation 
antigen, study day and participant: 
o 
IFN-𝛾 and/or TNF and/or IL-2. 
o 
IFN-𝛾 and/or TNF and/or IL-2 and/or IL-17 and/or IL-22. 
o 
IL-17. 
o 
IL-22. 
15.2.1. 
DERIVATIONS 
15.2.1.1. 
Positive (G,2,T,17,22) 
The ‘Positive (G,2,T,17,22)’ (%) result per participant, T cell type, stimulation antigen and visit is derived as: 
∑𝐶𝑒𝑙𝑙 𝑝𝑒𝑟𝑐𝑒𝑛𝑡𝑎𝑔𝑒𝑠 (%) 𝑤ℎ𝑒𝑟𝑒 𝐼𝐹𝑁−𝑔 𝑎𝑛𝑑/𝑜𝑟 𝐼𝐿−2  𝑎𝑛𝑑 / 𝑜𝑟 𝑇𝑁𝐹 𝑎𝑛𝑑 / 𝑜𝑟 𝐼𝐿-17 𝑎𝑛𝑑 /
 𝑜𝑟 𝐼𝐿-22 𝑎𝑟𝑒 𝑝𝑜𝑠𝑖𝑡𝑖𝑣𝑒. 
15.2.1.2. 
Positive (G,2,T) 
The ‘Positive (G,2,T)’ (%) result per participant, T cell type, stimulation antigen and visit is derived as: 
∑𝐶𝑒𝑙𝑙 𝑝𝑒𝑟𝑐𝑒𝑛𝑡𝑎𝑔𝑒𝑠 (%) 𝑤ℎ𝑒𝑟𝑒 𝐼𝐹𝑁−𝑔 𝑎𝑛𝑑/𝑜𝑟 𝐼𝐿−2  and / 𝑜𝑟 𝑇𝑁𝐹 𝑎𝑟𝑒 𝑝𝑜𝑠𝑖𝑡𝑖𝑣𝑒. 
15.2.1.3. 
Positive (17) 
The ‘Positive (17)’ (%) result per participant, stimulation antigen and visit for CD4 T cells is derived as: 
∑𝐶𝑒𝑙𝑙 𝑝𝑒𝑟𝑐𝑒𝑛𝑡𝑎𝑔𝑒𝑠 (%) 𝑤ℎ𝑒𝑟𝑒 𝐼𝐿−17  𝑖𝑠 𝑝𝑜𝑠𝑖𝑡𝑖𝑣𝑒. 
15.2.1.4. 
Positive (22) 
The ‘Positive (22)’ (%) result per participant, stimulation antigen and visit for CD4 cells is derived as: 
∑𝐶𝑒𝑙𝑙 𝑝𝑒𝑟𝑐𝑒𝑛𝑡𝑎𝑔𝑒𝑠 (%) 𝑤ℎ𝑒𝑟𝑒 𝐼𝐿−22  𝑖𝑠 𝑝𝑜𝑠𝑖𝑡𝑖𝑣𝑒. 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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15.2.2. 
RESPONDER STATUS 
The same method as for the PBMC ICS analysis will be used (refer to Section 15.1.2: Responder Status). 
Substitute ‘Positive (G,2)’ with each applicable derived cytokine combination.  
16. SAFETY OUTCOMES 
Summary and presentation of safety data will be based on the Safety analysis set, unless otherwise noted. 
16.1. ADVERSE EVENTS (AES) 
Adverse events (AEs) will be coded using Medical Dictionary for Regulatory Activities (MedDRA) Version 16.1. 
The following AE variables, as presented in the planned TLF shells, will be collected on the Adverse Event (AE) 
eCRF: 
• 
Verbatim term (eCRF). 
• 
Date started. 
• 
Date stopped. 
• 
Serious: 
o 
Yes. 
o 
No. 
• 
Severity (refer to Section 16.1.2.1: Severity). 
• 
Relationship to study vaccination (refer to Section 16.1.2.2: Relationship to Study Vaccination). 
• 
Outcome: 
o 
Resolved. 
o 
Resolved with sequelae. 
o 
Ongoing. 
o 
Death. 
• 
Did the AE cause the participant to discontinue from the study: 
o 
Yes. 
o 
No. 
• 
Was treatment given: 
o 
Yes. 
o 
No. 
• 
Is the AE of special interest: 
o 
Yes. 
o 
No. 
• 
System Organ Class (SOC). 
• 
Preferred Term (PT). 


 
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The following AE variables, as presented in the planned TLF shells, will be collected on the SAE Report eCRF: 
• 
Seriousness criterion: 
o 
Death. 
o 
Life-threatening. 
o 
Hospitalization. 
o 
Congenital anomaly/birth defect. 
o 
Involved persistent disability/incapacity. 
o 
Other important medical condition. 
o 
Prolonged hospitalization. 
16.1.1. 
DERIVATIONS 
Based on the aforementioned, the following variable, as presented in the planned TLF shells, will be derived: 
• 
Study day of the most recent vaccination: 
o 
Calculated relative to Day 0 vaccination (refer to Section 6.1: Reference Start Date and Study Day). 
• 
Relative days (start and stop): 
o 
Calculated relative to most recent vaccination (refer to Section 6.1: Reference Start Date and Study 
Day). 
• 
Dose at onset (most recent vaccination): 
o 
Refer to Section 16.1.8: Adverse Event (AE) Dose at Onset (Most Recent Vaccination). 
• 
Time of onset: 
o 
Derived based on the relative days (start) as either of the following categories: 
▪ 
Day 0 to Day 3. 
▪ 
Day 4 to Day 7. 
▪ 
Day 8 to Day 14. 
▪ 
>= Day 15. 
o 
For the overview of unsolicited AEs by severity, time of onset and relationship table, AEs with 
missing time to onset will be considered. 
16.1.2. 
ALL ADVERSE EVENTS (AES) 
The following tables are planned for presentation: 
• 
Overview of AEs: 
o 
Number and percentage (%) of participants in each of the following categories (participants with 
multiple events in each category are counted only once in each category): 
▪ 
Any AE. 
▪ 
Any solicited AE. 
▪ 
Local solicited AE. 
▪ 
Systemic solicited AE. 
▪ 
SAE. 
▪ 
SAE with outcome of death. 
▪ 
Related AE. 


 
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▪ 
Severe AE. 
▪ 
Related, severe AE. 
▪ 
AE leading to premature study discontinuation. 
▪ 
Adverse events of special interest (AESI). 
o 
Percentage (%) of participants will be calculated relative to the total number of participants in the 
Safety analysis set per treatment group and cohort. 
o 
95% CIs: 
▪ 
Calculated for single proportion using mid-p binomial option (Clopper-Pearson method with 
mid-p correction). 
o 
Number of events in each of the aforementioned categories (participants with multiple events in 
each category are counted multiple times in each category). 
o 
This output will be repeated for the Safety and Immunogenicity Cohort as well as the Correlates 
Cohort.  
All AE incidence tables will be sorted by: 
• 
SOC (descending frequency of total number of participants with events reported). 
• 
PT (descending frequency of total number of participants with events reported within each SOC). 
All AE incidence tables presented by PT only will be sorted by: 
• 
PT (descending frequency of total number of participants with events reported).  
The following incidence table will be presented by SOC and PT: 
• 
Incidence of AEs: 
o 
Number and percentage (%) of all participants with AEs summarized by SOC and PT. 
o 
Includes all AEs within the post-vaccination reporting windows (refer to Section 16.1.9: Adverse 
Event (AE) Post-vaccination Reporting Window for details on the derivation of post-vaccination 
reporting window). 
o 
This table will be repeated for the Safety and Immunogenicity Cohort. 
• 
Incidence of AEs by dose (refer to 16.1.8: Adverse Event (AE) Dose at Onset (Most Recent Vaccination)).  
The following incidence table is planned for presentation by PT only: 
• 
Incidence of AEs: 
The following listings are planned for presentation: 
• 
Solicited and unsolicited non-serious AEs.  
• 
AE comments.  
16.1.2.1. 
Severity 
Severity will be reported on the Adverse Events eCRF as mild, moderate or severe (increasing severity). 
Missing severities will not be regarded as ‘worst case’ severity. If any missing severity is present in the data 
transfer, the severity will remain missing and will be counted under the missing category in the by-severity 
tables. Participants with multiple events in each SOC/solicited AE category and PT are counted only once in 
each SOC/solicited AE category and PT at highest severity.  


 
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For the overview of unsolicited AEs by severity, time of onset and relationship table, AEs with missing severity 
will be considered. The participant will be counted in the missing category if the all severities are reported as 
missing for the relevant participant. 
The following incidence tables are planned for presentation by SOC and PT: 
• 
Incidence of AEs by highest severity.  
• 
Incidence of AEs by highest severity by dose (refer to 16.1.8: Adverse Event (AE) Dose at Onset (Most 
Recent Vaccination)).  
The following listing will be presented: 
• 
Adverse events of toxicity grade 3/4: 
o 
An AE of toxicity grade 3/4 is any AE for which the severity is indicated as severe. 
16.1.2.2. 
Relationship to Study Vaccination 
Relationship to study vaccination will be reported on the Adverse Events eCRF as definite, probable, possible, 
unlikely or not related (decreasing relationship). 
The relationship to study vaccination will be reported as either related or not related: 
• 
Related AE if relationship to study vaccination is reported as: 
o 
Definite. 
o 
Probable. 
o 
Possible. 
• 
Not related AE if relationship to study vaccination is reported as: 
o 
Unlikely. 
o 
Not related. 
Missing relationship to study vaccination will not be regarded as ‘worst case’ relationship to study vaccination. 
If any missing relationship to study vaccination is present in the data transfer, that relationship to study 
vaccination is to remain missing and will be counted in the missing category in the by-relationship tables. 
Participants with multiple events in each SOC/solicited AE category and PT are counted only once in each 
SOC/solicited AE category and PT at the strongest relationship. 
For the overview of unsolicited AEs by severity, time of onset and relationship table, AEs with missing 
relationship to study vaccine will be considered. The participant will be counted in the missing category if the 
all relationship to study vaccine is reported as missing for the relevant participant. 
The following incidence tables are planned for presentation by SOC and PT: 
• 
Incidence of related AEs.  
• 
Incidence of related AEs by dose (refer to 16.1.8: Adverse Event (AE) Dose at Onset (Most Recent 
Vaccination)).  
• 
Incidence of related AEs by highest severity.  
• 
Incidence of related AEs by highest severity and dose (refer to 16.1.8: Adverse Event (AE) Dose at Onset 
(Most Recent Vaccination)).  


 
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The following incidence tables are planned for presentation by PT only: 
• 
Incidence of related AEs.  
16.1.3. 
UNSOLICITED ADVERSE EVENTS (AES) 
The following overview table is planned for presentation: 
• 
Overview of unsolicited AEs by severity, time of onset and relationship: 
o 
Number and percentage (%) of participants in each of the following categories (participants with 
multiple events in each category are counted only once in each category, worst case/shortest time to 
onset): 
▪ 
Severity (refer to Section 16.1.2.1: Severity). 
▪ 
Time of onset (refer to Section 16.1.1: Derivations). 
▪ 
Relationship (refer to Section 16.1.2.2: Relationship to Study Vaccination). 
o 
Number of events in each of the aforementioned categories (participants with multiple events in 
each category are counted multiple times in each category). 
o 
Includes all unsolicited AEs, SAEs as well as AEs of special interest within the post-vaccination 
reporting window (refer to Section 16.1.9: Adverse Event (AE) Post-vaccination Reporting Window 
for details on the derivation of the post-vaccination reporting window). 
o 
95% CIs: 
▪ 
Calculated for single proportion using mid-p binomial option (Clopper-Pearson method with mid-
p correction). 
o 
This table will be repeated for the Safety and Immunogenicity Cohort as well as the Correlates 
Cohort.  
16.1.4. 
SOLICITED AND INJECTION SITE ADVERSE EVENTS (AES) 
In the database all AEs were reported as either solicited or unsolicited, however the database categorization 
will not be used to report AEs as either solicited or unsolicited. All AEs will be assigned as either solicited or 
unsolicited based on the PT (refer to APPENDIX 3: Solicited and Injection Site Adverse Event (AE)) and post-
vaccination reporting window (refer to Section 16.1.9: Adverse Event (AE) Post-vaccination Reporting Window 
for a description of the post-vaccination reporting windows). 
Local and systemic solicited AE incidence tables will be presented as follows: 
• 
Solicited AE category (local, systemic). 
• 
PT (descending frequency of total number of participants with events reported within each solicited AE 
category). 
The following incidence tables will be presented by PT only: 
• 
Incidence of solicited AEs.  
• 
Incidence of related solicited AEs.  
• 
Incidence of related solicited AEs by dose at onset (refer to Section 16.1.8: Adverse Event (AE) Dose at 
Onset (Most Recent Vaccination) for details on the derivation of dose at onset): 
o 
Number and percentage (%) of participants with AEs summarized by solicited AE category and PT. 


 
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o 
Includes all solicited AEs within the post-vaccination reporting window (refer to Section 16.1.9: 
Adverse Event (AE) Post-vaccination Reporting Window for details on the derivation of the post-
vaccination reporting window). 
o 
BCG treatment group will only include Dose 1 at onset AEs. 
• 
Incidence of solicited AEs per cohort and dose at onset (refer to Section 16.1.8: Adverse Event (AE) Dose 
at Onset (Most Recent Vaccination) for details on the derivation of dose at onset): 
o 
Number and percentage (%) of participants with AEs summarized by solicited AE category and PT. 
o 
Includes all solicited AEs within the post-vaccination reporting window (refer to Section 16.1.9: 
Adverse Event (AE) Post-vaccination Reporting Window for details on the derivation of the post-
vaccination reporting window). 
o 
BCG treatment group will only include Dose 1 at onset AEs. 
• 
Incidence of local solicited AEs by severity and dose at onset: 
o 
Number and percentage (%) of participants with AEs summarized by solicited AE category and PT. 
o 
Includes all local solicited AEs within the post-vaccination reporting window (refer to Section 16.1.9: 
Adverse Event (AE) Post-vaccination Reporting Window for details on the derivation of the post-
vaccination reporting window). 
o 
BCG treatment group will only include Dose 1 at onset AEs. 
o 
This table will be repeated for systemic solicited AEs and related systemic solicited AEs. 
16.1.5. 
SERIOUS ADVERSE EVENTS (SAES) 
An SAE is any AE for which serious event is indicated as ‘yes’ on the Adverse Event eCRF. 
The following listing is planned for presentation by treatment group: 
• 
SAEs. 
• 
SAEs with an outcome of death.  
The following incidence table is planned for presentation by SOC and PT: 
• 
Incidence of serious adverse events by dose. 
• 
Incidence of serious adverse events with outcome of death.   
16.1.6. 
ADVERSE EVENTS OF SPECIAL INTEREST (AESI) 
An AESI is any AE for which AE of special interest is indicated as ‘yes’ on the Adverse Events eCRF. Refer to 
APPENDIX 4: Adverse Events of Special Interest (AESIs) for the list of AEs considered to be of special interest. 
The following listing is planned for presentation by treatment group: 
• 
Adverse events of special interest (AESI). 
16.1.7. 
ADVERSE EVENTS LEADING TO PREMATURE STUDY DISCONTINUATION 
An AE leading to premature study discontinuation is any AE for which did the AE cause the participant to 
discontinue from the study recorded as ‘yes’ 


 
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The following listing is planned for presentation by treatment group: 
• 
Adverse events leading to premature study discontinuation. 
16.1.8. 
ADVERSE EVENT (AE) DOSE AT ONSET (MOST RECENT VACCINATION) 
A dose at onset will be assigned for all AEs and is defined as the most recent dose prior to the onset of the AE: 
• 
Post any dose (combination of the AEs starting after the Day 0 and Day 56 vaccinations). 
• 
Post-dose 1 (AEs starting after the Day 0 vaccination). 
• 
Post-dose 2 (AEs staring after the Day 56 vaccination): 
o 
Only applicable to H4:IC31 and Placebo treatment groups. 
The assignment of the dose at onset will be based on the following algorithm: 
• 
Calculate the difference in the AE start date (as collected on the Adverse Events eCRF) and all available 
study vaccination dates (as collected on the Exposure Day 0 and Exposure Day 56 eCRFs): 
o 
Days since dose x: (AE start date – study vaccination dose x date [where x= 1 or 2]). 
• 
From the previous calculation, only consider those doses where the days since dose x is >=0 and 
determine the dose (1 or 2) for which the days since dose x (as calculated in the previous step) is a 
minimum. 
• 
Assign the dose selected in the previous step as the dose at onset as the specific dose determined in the 
previous calculation. 
16.1.9. 
ADVERSE EVENT (AE) POST-VACCINATION REPORTING WINDOW 
All SAEs and AESIs will be collected through database lock. 
All unsolicited AEs will be collected during the 28-day reporting window following each study vaccination. The 
reporting window is defined in terms of the relevant scheduled study visit coinciding with the 28-day window 
following the relevant dose. 
Solicited systemic adverse events will be collected during the 7-day reporting window following each study 
vaccination (diary cards will be used for 7 days following each study vaccination for the Safety and 
Immunogenicity Cohort only).  
Solicited and unsolicited local injection site reaction adverse events will be collected: 
• 
BCG treatment group: 
o 
84-day reporting window following study vaccination. 
• 
H4:IC31 and Placebo treatment groups: 
o 
28-day reporting window following each vaccination. 


 
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Refer to Table 4: General Adverse Event (AE) Post-vaccination Reporting Windows below for the relevant post-
vaccination reporting windows as defined in terms of scheduled study visits: 
Table 4: General Adverse Event (AE) Post-vaccination Reporting Windows 
Adverse Event 
Type 
Treatment 
Group 
Dose at 
Onset 
Post-vaccination Reporting Window 
Scheduled Study 
Visit 
Protocol Specified Visit Window 
Unsolicited AE 
All 
Day 0 
Study Day 28 
28 days following Study Day 0 
H4: IC31 
Placebo 
Day 56 
Study Day 84 
28 days following Study Day 56 
Local Solicited 
AEs 
BCG 
Day 0 
Study Day 84 
84 days following Study Day 0 
H4: IC31 
Placebo 
Day 0 
Study Day 28 
28 days following Study Day 0 
H4: IC31 
Placebo 
Day 56 
Study Day 84 
28 days following Study Day 56 
Systemic 
Solicited AEs 
All 
Day 0 
Study Day 7 
7 days  following Study Day 0 
H4: IC31 
Placebo 
Day 56 
Study Day 63 
7 days following Study Day 56 
SAEs and AEs of 
special interest 
All 
All 
- 
Through database lock 
If the relevant scheduled study visit occurred within the protocol specified visit window, all AEs with start date 
through the relevant scheduled study visit will be included in the reporting period. If the relevant scheduled 
study visit occurred outside the protocol specified visit window, only include AEs with start date through either 
7, 28 or 84 days will be included. 
16.2. INJECTION SITE REACTION ASSESSMENT 
Injection site reaction assessments will be presented for the Safety analysis set.  
The following injection site reaction assessment listing is planned for presentation: 
• 
Injection site reaction assessment.  
The following injection site reaction variables will be collected throughout the study on the eCRFs: 
• 
Date and time of assessment.  
• 
Injection site reaction assessment: 
o 
Pain.  
o 
Redness.  
o 
Swelling.  
o 
Ulceration.  
o 
Drainage.  
o 
Abnormality photo taken.  
o 
Axillary lymphadenopathy.  


 
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• 
Present: 
o 
Yes: 
▪ 
Measurement (mm) for relevant assessments.  
o 
No.  
16.2.1. 
DERIVATIONS 
The following variables, as presented in the planned TLF shells, will be derived: 
• 
Any time point: 
o 
Per study vaccination and injection site reaction the worst case result is selected as the any 
timepoint assessment.  
o 
Worst case assessment is defined as the largest measurement (mm) for assessments with 
measurements or the presence of the reaction if o measurements are available.  
• 
Overall time point: 
o 
Per injection site assessment the worst case result is selected as the overall assessment.  
o 
Worst case assessment is defined as for the any timepoint assessment per study vaccination.  
16.3. LABORATORY EVALUATIONS 
The following tables are planned for presentation (based on the Safety analysis set for the Safety and 
Immunogenicity Cohort): 
• 
Quantitative laboratory results and change from Baseline: 
o 
Presented by means of default summary statistics for quantitative variables. 
o 
Change from Baseline will be presented as well. 
• 
Shift in laboratory grades from Baseline: 
o 
Shift will be presented in terms of the number of participants with an increased toxicity grade from 
the Baseline result (refer to APPENDIX 7: Toxicity Grading Criteria): 
▪ 
0 grades (no change in grade compared to Baseline or grade decreased).  
▪ 
1 grade. 
▪ 
2 grades.  
▪ 
3 grades.  
▪ 
4 grades (only applicable to certain tests).  
o 
N1 is defined as the total number of participants with an assessment available at both Baseline and 
post-baseline time points.  
o 
Due to the fact that the toxicity grading for Urine Erythrocytes is “Normal”, “Grade 1” or “Grade 
2/3/4”, it is not possible to determine if there was an increase. Therefore Urine Erythrocytes will not 
be included in this table. The toxicity grading will be displayed in the listing.  
The following listings are planned for presentation (based on the Randomized analysis set): 
• 
Quantitative laboratory results.  
• 
Qualitative laboratory results.  
• 
Laboratory reference ranges by system and test (no analysis set applicable to this listing).  


 
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Refer to APPENDIX 5: Laboratory Assessments for a list of laboratory assessments.  
The following laboratory variables, as presented in the planned TLF shells, will be collected during the study on 
the Hematology (HEMA), Chemistry (CHEM), Urinalysis (URIN), Pregnancy Test (PREG) and HIV Tests (HIVT) 
eCRFs: 
• 
Date of sample collection.  
• 
Time of sample collection (only applicable to pregnancy test results).  
• 
Test.  
• 
Reported results and unit.  
All conversion of original to standard units will be completed by Data Management within the database prior to 
transfer to Biostatistics. All laboratory data will be summarized in the standard unit.  
16.3.1. 
TOXICITY GRADING CRITERIA 
The toxicity grades will be assigned based on the protocol-specific criteria specified in APPENDIX 7: Toxicity 
Grading Criteria. For some assessments grade 3 or 4 cannot be assigned programmatically, but rather are based 
on clinical judgment, therefore they are not assigned for the purpose of analysis.  
Refer to APPENDIX 5: Laboratory Assessments for a list of tests for which toxicity grades will be 
programmatically assigned.  
16.4. VITAL SIGNS 
Vital signs will be presented for the Safety analysis set. 
The following tables are planned for presentation: 
• 
Vital signs and change from pre-vaccination: 
o 
Presented by means of default summary statistics for quantitative variables. 
o 
Change from pre-vaccination will be presented as well. 
• 
Shift in vital signs toxicity grades from pre-vaccination (Study Day 0 or Study Day 56 [not applicable to 
BCG]): 
o 
Shift will be presented in terms of the number of participants with an increased toxicity grade from 
the pre-vaccination results (refer to): 
▪ 
0 grades (no change in grade compared to Baseline or grade decreased).  
▪ 
1 grade.  
▪ 
2 grades.  
▪ 
3 grades. 
o 
N1 is defined as the total number of participants with an assessment available at both the relevant 
pre- and post-vaccination time points who received the applicable study vaccine dose.  


 
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The following listing is planned for presentation (based on the Randomized analysis set): 
• 
Vital signs results.  
The following vital signs variables, as presented in the planned TLF shells, will be collected on the Vital Signs, 
Vital Signs - Pre-vaccination and Vital Signs - Post-vaccination eCRFs: 
• 
Result and unit for each of the following test: 
o 
Axillary temperature (˚C). 
o 
Pulse rate (beats per minute). 
o 
Systolic and diastolic blood pressure (mmHg). 
16.4.1. 
TOXICITY GRADING CRITERIA 
The toxicity grades will be assigned based on the protocol-specific criteria specified in APPENDIX 7: Toxicity 
Grading Criteria. Grade 4 cannot be assigned programmatically but rather is based on clinical judgment, 
therefore not assigned for the purpose of analysis.  
Refer to APPENDIX 6: Vital Signs Assessments for a list of tests for which toxicity grades will be 
programmatically assigned.  
16.4.2. 
PRE-VACCINATION AND POST-VACCINATION TIME POINTS 
Refer to Table 4: Vital Signs Pre-and Post-vaccination Time Points below for details on the relevant pre-and 
post-vaccination time point for the vital signs assessments.  
Study Vaccination 
Pre-vaccination Time Point 
Post-vaccination Time Point 
Dose 1 
Study Day 0 (Pre-vaccination) 
Study Day 0 (Post-vaccination) 
Dose 2 (not applicable to BCG) 
Study Day 56 (Pre-vaccination) 
Study Day 56 (Post-vaccination)  
16.5. PHYSICAL EXAMINATION  
Physical examination data will be presented for the Randomized analysis set. 
The following listing is planned for presentation: 
• 
Abnormal physical examination findings: 
o 
Only include physical examination results where the result is indicated as abnormal. 
The following physical examination variable, as presented in the planned TLF shells, will be collected on the 
Physical Examination eCRF: 
• 
Date of assessment. 
• 
Verbatim term (and abnormality description if applicable). 
 
 


 
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16.6. PREGNANCIES 
Pregnancies will be presented for the Safety analysis set. 
The following listing is planned for presentation: 
• 
Reported pregnancies. 
The following pregnancy variables, as presented in the planned TLF shells, will be collected on the Pregnancy 
Notification eCRF: 
• 
Pregnancy test: 
o 
Date of test. 
o 
Type: 
▪ 
Serum. 
▪ 
Urine. 
o 
Result: 
▪ 
Positive. 
▪ 
Negative. 
• 
Current pregnancy information: 
o 
Estimated conception date. 
o 
Estimated delivery date. 
• 
Current pregnancy outcome: 
o 
Date of delivery/outcome. 
o 
Outcome: 
▪ 
Normal. 
▪ 
Abnormal. 
• 
Health status of mother. 
• 
Health status of child. 
• 
Additional information. 
16.6.1. 
DERIVATIONS 
Based on the aforementioned the following variable, as presented in the planned TLF shells, will be derived: 
• 
Study day of the most recent vaccination: 
o 
Calculated relative to Day 0 vaccination (refer to Section 6.1: Reference Start Date and Study Day). 
• 
Relative day: 
o 
Calculated relative to most recent vaccination (refer to Section 6.1: Reference Start Date and Study 
Day). 
 
 


 
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17. DATA NOT SUMMARIZED OR PRESENTED 
The following eCRF data will not be presented: 
• 
Immediately Reportable Event eCRF:  
o 
Immediately reportable events would be reported as part of the relevant domain’s analysis.  


 
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Page 72 of 91 
18. REFERENCES 
Aeras C-040-404 Study Protocol Version 5.0, dated 25 July 2016. 
Aeras C-040-404 Electronic Case Report Form Version 6.0, dated 22APR2015. 
Aeras C-040-404 Primary Analysis Note to File dated 25 April 2016.  
 
 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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APPENDIX 1. PROGRAMMING CONVENTIONS FOR OUTPUTS 
Refer to the latest version of the relevant TLF shells. 
STUDY VISITS 
All study visits will be indicated as Visit in the relevant outputs (TLFs). The visits will be presented in the 
following order: 
• 
Screening. 
• 
Day x, where x = 0, 3, 7, 28, 56, 63, 70, 84. 
• 
Month x, where x = 6, 9, 12, 15, 18, 21, 24, 27, 30. 
• 
Any Time Point (if relevant). 
FOOTNOTES 
Footnotes will be ordered as follows: 
• 
Non-standard abbreviations, separated by a full stop. 
• 
n = Number of participants… N = Total number of participants… % = Percentage of participants. 
• 
Definitions. 
• 
Footnotes pertaining to statistical methodology. 
• 
If coding is presented, the version of the coding dictionary used. 
• 
Study-specific footnotes to clarify data points within the specific presentation. 
OUTPUT HEADERS 
Refer to the relevant set of TLF shells for the specific output header format. 
OUTPUT FOOTERS 
Refer to the relevant set of TLF shells for the specific output footer format. 
DECIMAL PRECISION 
All values will be rounded using the SAS® function ROUND, as the last step prior to presentation. All computed 
percentages (%) will be presented using one decimal place. If the original data has N decimal places (as derived 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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from the raw data), then the summary statistics for quantitative data are to contain the following number of 
decimal places (with a maximum of 3 places): 
• 
Minimum and maximum: N. 
• 
Mean and median: (N+1). 
• 
Standard deviation: (N+2). 
All p-values will be presented with 4 decimal places, if less than 0.0001 (after rounding to 4 decimal places) the 
p-value will be displayed as <0.0001. Odds ratios will be presented to 3 decimals and CIs will be presented to 2 
decimals. 
DATES & TIMES 
Dates will be displayed as DDMMMYYYY. Times will be displayed as HH:MM (24-hour clock). 
SPELLING FORMAT 
English US. 
LISTINGS 
All listings will be sorted primarily by treatment group, in the order specified as applicable to the specific listing. 
PRESENTATION NOTES 
• 
In all listings, participants will be identified by means of the 16-digit participant identifier. 
• 
Output margins should be top 1.5 inch and 1 inch on left, right and bottom. 
• 
Each TLF will be produced as a separate stand-alone document. 
• 
If separate presentations are required for analysis set or subgroups then these should be presented as 
stand-alone. 
• 
All spelling is to adhere to English (US). 
• 
All TLFs will be presented in a landscape format, as far as is feasible. 
• 
The content of all tables and listings will be uppercase (excluding titles, column headers and footnotes), 
that is the ‘Reported Term’ in the medical conditions listing, for example. 
• 
For all AE/CM/MH listings all coded terms will not be presented in uppercase, but rather as coded 
(sentence case) for differentiation from verbatim terms. 
 
 


 
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• 
The margin, page size and line size specifications as stipulated below will be used for the presentation of 
all TLFs: 
 
Landscape 
Portrait 
Paper Size 
U.S. Letter (8.5 x 11 inch) 
U.S. Letter (8.5 x 11 inch) 
Margins (Inches): 
Top  
Bottom 
Left 
Right 
 
1.25 
1 
1 
1 
 
1 
1 
1.25 
1 
Header (Inches) 
0.5 
0.5 
Footer (Inches) 
0.5 
0.5 
SAS® Specifications: 
PAGESIZE 
 
LINESIZE 
 
46 (8 point size, Courier) 
41 (9 point size, Courier) 
134 (8 point size, Courier) 
119 (9 point size, Courier) 
 
67 (8 point size, Courier) 
60 (9 point size, Courier) 
93 (8 point size, Courier) 
82 (9 point size, Courier) 
 
 


 
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APPENDIX 2. PARTIAL DATE CONVENTIONS 
Imputation will only be performed for partially or completely missing AE start dates. Imputed AE start dates will 
not be presented in the listings and are only used to assign AEs to the relevant post-vaccination reporting 
window. 
AE Start Date 
Criterion 
Action 
Day 
Month 
Year 
Missing 
Known 
Known 
AE start month and year are the 
same as the month and year of 
only one study vaccination (Day 
0 or Day 56) 
Assign the missing day as the day 
of the relevant study vaccination 
in the coinciding month and year 
AE start month and year are the 
same as the month and year of 
more than one study vaccination 
(Day 0 or Day 56) 
Assign the missing day as the day 
of the earliest study vaccination 
in the coinciding month and year 
AE start month and year are not 
the same as the month and year 
of any of the study vaccinations 
(Day 0 or Day 56) and known to 
be in between study 
vaccinations 
Assign the missing day as the first 
day of the month 
AE start month and year are not 
the same as the month and year 
of any of the study vaccinations 
(Day 0 or Day 56) and known to 
be prior to all study vaccinations 
Assign the missing day as the last 
day of the month 
AE start month and year are not 
the same as the month and year 
of any of the study vaccinations 
(Day 0 or Day 56) and known to 
be after all study vaccinations 
Assign the missing day as the first 
day of the month 
Missing 
Missing 
Known 
AE start year is the same as the 
year of only one study 
vaccination (Day 0 or Day 56) 
Assign the missing day and 
month as the day and month of 
the relevant study vaccination in 
the coinciding year 
AE start year is the same as the 
year of more than one study 
vaccination (Day 0 or Day 56) 
Assign the missing day and 
month as the day and month of 
the earliest study vaccination in 
the coinciding year 
AE start year is not the same as 
the year of any of the study 
vaccinations (Day 0 or Day 56) 
and known to be prior to all 
study vaccinations 
Assign the missing day and 
month as 31 December 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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AE Start Date 
Criterion 
Action 
Day 
Month 
Year 
AE start year is not the same as 
the year of any of the study 
vaccinations (Day 0 or Day 56) 
and known to be after all study 
vaccinations 
Assign the missing day and 
month as 01 January 
Missing 
Missing 
Missing 
- 
Assign the missing date as the 
date of Dose 1 of study 
vaccination 
 
 
 


 
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APPENDIX 3. SOLICITED AND INJECTION SITE ADVERSE EVENT (AE) 
PREFERRED TERMS AND CODES 
Relevant 
Treatment 
Group 
Solicited or 
Injection Site AE 
Solicited AE 
Category 
Preferred Term 
Preferred 
Term Code 
Placebo 
H4:IC31  
BCG 
Solicited AE 
Systemic 
Pyrexia 
10037660 
Myalgia 
10028411 
Arthralgia 
10003239 
Fatigue 
10016256 
Headache 
10019211 
Decreased appetite 
10061428 
Urticaria 
10046735 
Chills 
10008531 
Lymphadenopathy 
10025197 
BCG 
Injection site AE 
Local 
Vaccination site pain 
10068879 
Vaccination site erythema 
10059079 
Vaccination site swelling 
10069620 
Vaccination site ulcer 
10069621 
Vaccination site discharge 
10069560 
Placebo 
H4:IC31 
Injection site AE 
Local 
Injection site pain 
10022086 
Injection site erythema 
10022061 
Injection site swelling 
10053425 
Injection site ulcer 
10022105 
Injection site discharge 
10065600 
 
 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 79 of 91 
APPENDIX 4. ADVERSE EVENTS OF SPECIAL INTEREST (AESIS) 
Events, which are potentially immune, mediated are AESIs and include the following: 
Acute disseminated encephalomyelitis (ADEM) 
Myelitis/Transverse Myelitis 
Addison's Disease 
Myocarditis 
Anti-neutrophil Cytoplasmic Antibody (ANCA)-associated Vasculitis 
Nephritis 
Ankylosing Spondylitis 
Optic neuritis 
Anti-phospholipid Syndrome 
Pericarditis 
Autoimmune Bullous Skin Diseases 
Polymyalgia Rheumatica 
Autoimmune Hemolytic Anemia 
Polymyositis 
Autoimmune Hepatitis 
Primary Biliary Cirrhosis 
Autoimmune Thrombotic/Thromboembolic Conditions 
Primary Sclerosing Cholangitis 
Basedow’s Disease 
Psoriasis 
Behcet’s Syndrome 
Psoriatic Arthritis 
Bell's Palsy 
Raynaud's Phenomenon 
Carditis 
Rheumatoid Arthritis 
Celiac Disease 
Sarcoidosis 
Crohn’s Disease 
Scleroderma 
Cutaneous Lupus 
Sjogren’s Syndrome 
Demyelinating Disease 
Spondylo-arthropathy 
Dermatomyositis 
Stevens-Johnson Syndrome 
Diabetes Mellitus, Insulin Dependent (IDDM) 
Systemic Lupus Erythematosus 
Erythema Nodosum 
Temporal Arteritis 
Glomerulonephritis 
Thyroiditis 
Guillain Barre Syndrome 
Tolosa-Hunt Syndrome 
Grave’s Disease 
Ulcerative Colitis 
Idiopathic Thrombocytopenic Purpura (ITP) 
Ulcerative Proctitis 
Inflammatory Bowel Disease (non-specific) 
Uveitis 
Juvenile Rheumatoid Arthritis 
Vasculitis 
Mixed Connective Tissue Disease 
Vitiligo 
Multiple Sclerosis 
Wegener’s Granulomatosis 
Myasthenia Gravis 
 
 
 


 
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APPENDIX 5. LABORATORY ASSESSMENTS 
System 
   Test 
Reported Unit 
Conversion 
Factor 
SI Unit 
Toxicity Abnormality* 
Hematology 
   Basophils 
10^9/L 
1 
10^9/L 
- 
   Eosinophils 
10^9/L 
1 
10^9/L 
- 
   Hematocrit 
Fraction of one 
1 
Fraction of % 
- 
L/L 
1 
Fraction of % 
- 
   Hemoglobin 
g/dL 
10 
g/L 
Hemoglobin -decreased 
   Lymphocytes 
10^9/L 
1 
10^9/L 
- 
   Monocytes 
10^9/L 
1 
10^9/L 
- 
   Neutrophils 
10^9/L 
1 
10^9/L 
- 
   Platelet count 
10^9/L 
1 
10^9/L 
Platelets - decreased 
   White blood cell  
  (WBC) count 
10^9/L 
1 
10^9/L 
WBC - decreased 
WBC - increased 
Chemistry 
   Alkaline 
phosphatase 
U/L 
1 
U/L 
Alkaline phosphatase - 
increased 
   ALT 
U/L 
1 
U/L 
ALT (SGPT) - increased 
   AST 
U/L 
1 
U/L 
AST (SGOT) - increased 
   Blood urea  
   nitrogen 
mg/dL 
0.357 
mmol/L 
BUN - increased 
mmol/L 
1 
mmol/L 
   Creatinine 
𝜇mol/L 
1 
𝜇mol/L 
Creatinine - increased 
   Total bilirubin 
𝜇mol/L 
1 
𝜇mol/L 
Total bilirubin - increased 
Urinalysis 
   Blood 
No standardization performed for the urinalysis 
laboratory assessments 
- 
   Glucose 
- 
   Protein 
Proteinuria, random collection 
   Leucocytes 
- 
   Erythrocytes 
Hematuria 
   Epithelial cells 
- 
*Refer to APPENDIX 7: Toxicity Grading Criteria. 
 


 
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APPENDIX 6. VITAL SIGNS ASSESSMENTS 
Test 
Unit 
Toxicity 
Abnormality* 
Axillary body temperature 
˚C 
Fever 
Pulse rate 
bpm 
- 
Blood pressure: Diastolic 
mmHg 
Hypertension 
Blood pressure: Systolic 
mmHg 
Hypertension 
Height 
cm 
- 
Weight 
kg 
- 
BMI 
kg/m2 
- 
*Refer to APPENDIX 7: Toxicity Grading Criteria. 
 
 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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APPENDIX 7. TOXICITY GRADING CRITERIA 
ESTIMATING SEVERITY GRADE 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Clinical 
adverse 
event NOT 
identified 
elsewhere 
in this AE 
Grading 
Table 
Symptoms 
causing no or 
minimal 
interference 
with usual social 
& functional 
activities 
Symptoms causing 
greater than 
minimal 
interference with 
usual social & 
functional activities 
Symptoms 
causing 
inability to 
perform usual 
social & 
functional 
activities  
Symptoms causing 
inability to perform 
basic self-care functions 
OR Medical or operative 
intervention indicated 
to prevent permanent 
impairment, persistent 
disability, or death 
CARDIOVASCULAR 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Hypertension 
Adult > 17 
years  
(with repeat 
testing at 
same visit)  
140 – 159 
mmHg systolic 
OR  
90 – 99 mmHg 
diastolic  
160 – 179 mmHg 
systolic OR  
100 – 109 mmHg 
diastolic  
≥ 180 mmHg 
systolic OR  
≥ 110 mmHg 
diastolic  
Life-threatening 
consequences (e.g., 
malignant 
hypertension) OR 
Hospitalization 
indicated (other than 
emergency room visit)  
Hypertension 
Pediatric ≤ 
17 years  
(with repeat 
testing at 
same visit)  
NA  
91st – 94th 
percentile 
adjusted for age, 
height, and 
gender (systolic 
and/or diastolic)  
146 - 168 mmHg 
systolic  
OR  
91 - 102 mmHg 
diastolic  
≥ 95th percentile 
adjusted for age, 
height, and gender 
(systolic and/or 
diastolic)  
≥168 mmHg  
systolic  
OR  
≥102 mmHg  
diastolic  
Life-threatening 
consequences (e.g., 
malignant 
hypertension) OR 
Hospitalization 
indicated (other than 
emergency room visit) 
Hypotension  
NA  
Symptomatic, 
corrected with 
oral fluid 
replacement  
Symptomatic, IV 
fluids indicated  
Shock requiring use of 
vasopressors or 
mechanical assistance 
to maintain blood 
pressure  


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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SKIN – DERMATOLOGICAL 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Cutaneous 
reaction – rash  
Localized 
macular rash  
Diffuse macular, 
maculopapular, or 
morbilliform rash 
OR Target lesions  
Diffuse macular, 
maculopapular, or 
morbilliform rash 
with vesicles or 
limited number of 
bullae OR 
Superficial 
ulcerations of 
mucous 
membrane limited 
to one site  
Extensive or 
generalized bullous 
lesions OR Stevens-
Johnson syndrome 
OR Ulceration of 
mucous membrane 
involving two or more 
distinct mucosal sites 
OR Toxic epidermal 
necrolysis (TEN)  
Pruritus 
(itching – no 
skin lesions)  
(See also 
Injection Site 
Reactions: 
Pruritus 
associated 
with injection)  
Itching 
causing no or 
minimal 
interference 
with usual 
social & 
functional 
activities  
Itching causing 
greater than 
minimal 
interference with 
usual social & 
functional 
activities  
Itching causing 
inability to 
perform usual 
social & functional 
activities  
NA  
INFECTION 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Infection (any 
other than HIV 
infection)  
Localized, no 
systemic 
antimicrobial 
treatment 
indicated 
AND 
Symptoms 
causing no or 
minimal 
interference 
with usual 
social & 
functional 
activities  
Systemic 
antimicrobial 
treatment 
indicated OR 
Symptoms causing 
greater than 
minimal 
interference with 
usual social & 
functional 
activities  
Systemic antimicrobial 
treatment indicated 
AND Symptoms 
causing inability to 
perform usual social & 
functional activities 
OR Operative 
intervention (other 
than simple incision 
and drainage) 
indicated  
Life-threatening 
consequences 
(e.g., septic 
shock)  
 
 


 
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STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
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SYSTEMIC 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Acute systemic 
allergic reaction  
 
Localized urticaria 
(wheals) with no 
medical 
intervention 
indicated  
Localized urticaria 
with medical 
intervention 
indicated OR Mild 
angioedema with 
no medical 
intervention 
indicated  
Generalized 
urticaria OR 
Angioedema with 
medical 
intervention 
indicated OR 
Symptomatic 
mild 
bronchospasm  
Acute 
anaphylaxis OR 
Life-threatening 
bronchospasm 
OR laryngeal 
edema  
Chills  
Symptoms 
causing no or 
minimal 
interference with 
usual social & 
functional 
activities  
Symptoms 
causing greater 
than minimal 
interference with 
usual social & 
functional 
activities  
Symptoms 
causing inability 
to perform usual 
social & 
functional 
activities  
NA  
Fatigue  
Malaise  
Symptoms 
causing no or 
minimal 
interference with 
usual social & 
functional 
activities  
Symptoms 
causing greater 
than minimal 
interference with 
usual social & 
functional 
activities  
Symptoms 
causing inability 
to perform usual 
social & 
functional 
activities  
Incapacitating 
fatigue/ malaise 
symptoms 
causing inability 
to perform 
basic self-care 
functions  
Fever (axillary)  
38.0 – 38.4°C  
100.4 – 101.1°F  
38.5 - 40°C  
101.2 - 104°F  
>40°C  
>104°F  
NA  
Pain (indicate body 
site)  
DO NOT use for 
pain due to 
injection (See 
Injection Site 
Reactions: 
Injection site pain)  
See also Headache, 
Arthralgia, and 
Myalgia  
Pain causing no 
or minimal 
interference with 
usual social & 
functional 
activities  
Pain causing 
greater than 
minimal 
interference with 
usual social & 
functional 
activities  
Pain causing 
inability to 
perform usual 
social & 
functional 
activities  
Disabling pain 
causing inability 
to perform 
basic self-care 
functions OR 
Hospitalization 
(other than 
emergency 
room visit) 
indicated 


 
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Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Unintentional 
weight loss  
NA  
5 – 9% loss in 
body weight from 
baseline  
10 – 19% loss in 
body weight from 
baseline  
≥ 20% loss in 
body weight 
from baseline 
OR Aggressive 
intervention 
indicated [e.g., 
tube feeding or 
total parenteral 
nutrition (TPN)]  
INJECTION SITE REACTIONS 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Injection site 
pain (pain 
without 
touching) Or 
Tenderness 
(pain when 
area is 
touched)  
Pain/tenderness 
causing no or 
minimal limitation 
of use of limb  
Pain/tenderness 
limiting use of limb 
OR 
Pain/tenderness 
causing greater 
than minimal 
interference with 
usual social & 
functional 
activities  
Pain/tenderness 
causing inability 
to perform usual 
social & 
functional 
activities  
Pain/tenderness 
causing inability to 
perform basic self-
care function OR 
Hospitalization 
(other than 
emergency room 
visit) indicated for 
management of 
pain/tenderness  
Injection site 
reaction 
(localized)  
 
Adult > 15 
years  
Erythema OR 
Induration of 5x5 
cm – 9x9 cm (or 25 
cm2 – 81cm2)  
Erythema OR 
Induration OR 
Edema > 9 cm any 
diameter (or > 81 
cm2)  
Ulceration OR 
Secondary 
infection OR 
Phlebitis OR 
Sterile abscess 
OR Drainage  
Necrosis (involving 
dermis and deeper 
tissue)  
Injection site 
reaction 
(localized)  
 
Pediatric ≤ 15 
years  
Erythema OR 
Induration OR 
Edema present but 
≤ 2.5 cm diameter  
Erythema OR 
Induration OR 
Edema > 2.5 cm 
diameter but < 
50% surface area 
of the extremity 
segment (e.g., 
upper arm/thigh)  
Erythema OR 
Induration OR 
Edema involving 
≥ 50% surface 
area of the 
extremity 
segment (e.g., 
upper arm/thigh) 
OR Ulceration OR 
Secondary 
infection OR 
Phlebitis OR 
Sterile abscess 
OR Drainage  
Necrosis (involving 
dermis and deeper 
tissue) 


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
The contents of this document are confidential and proprietary to Quintiles Transnational Corp. Unauthorized use, disclosure or 
reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 86 of 91 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Pruritus 
associated 
with injection  
See also Skin: 
Pruritus 
(itching - no 
skin lesions)  
Itching localized to 
injection site AND 
Relieved 
spontaneously or 
with < 48 hours 
treatment  
Itching beyond the 
injection site but 
not generalized OR 
Itching localized to 
injection site 
requiring ≥ 48 
hours treatment  
Generalized 
itching causing 
inability to 
perform usual 
social & 
functional 
activities  
NA  
GASTROINTESTINAL 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Anorexia* 
Loss of appetite 
without decreased 
oral intake  
Loss of appetite 
associated with 
decreased oral 
intake without 
significant weight 
loss  
Loss of appetite 
associated with 
significant weight 
loss  
Life-threatening 
consequences OR 
Aggressive 
intervention 
indicated [e.g., 
tube feeding or 
total parenteral 
nutrition (TPN)]  
Diarrhea  
Transient or 
intermittent 
episodes of 
unformed stools OR 
Increase of ≤ 3 
stools over baseline 
per 24-hour period  
Persistent episodes 
of unformed to 
watery stools OR 
Increase of 4 – 6 
stools over baseline 
per 24-hour period  
Bloody diarrhea OR 
Increase of ≥ 7 
stools per 24-hour 
period OR IV fluid 
replacement 
indicated  
Life-threatening 
consequences 
(e.g., hypotensive 
shock)  
Nausea  
Transient (< 24 
hours) or 
intermittent nausea 
with no or minimal 
interference with 
oral intake  
Persistent nausea 
resulting in 
decreased oral 
intake for 24 – 48 
hours  
Persistent nausea 
resulting in minimal 
oral intake for > 48 
hours OR 
Aggressive 
rehydration 
indicated (e.g., IV 
fluids)  
Life-threatening 
consequences 
(e.g., hypotensive 
shock)  
Vomiting  
Transient or 
intermittent 
vomiting with no or 
minimal 
interference with 
oral intake  
Frequent episodes 
of vomiting with no 
or mild dehydration  
Persistent vomiting 
resulting in 
orthostatic 
hypotension OR 
Aggressive 
rehydration 
indicated (e.g., IV 
fluids)  
Life-threatening 
consequences 
(e.g., hypotensive 
shock)  


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
The contents of this document are confidential and proprietary to Quintiles Transnational Corp. Unauthorized use, disclosure or 
reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 87 of 91 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Headache  
Symptoms causing 
no or minimal 
interference with 
usual social & 
functional activities  
Symptoms causing 
greater than 
minimal 
interference with 
usual social & 
functional activities  
Symptoms causing 
inability to perform 
usual social & 
functional activities  
Symptoms 
causing inability 
to perform basic 
self-care 
functions OR 
Hospitalization 
indicated (other 
than emergency 
room visit) OR 
Headache with 
significant 
impairment of 
alertness or other 
neurologic 
function  
Insomnia  
NA  
Difficulty sleeping 
causing greater 
than minimal 
interference with 
usual social & 
functional activities  
Difficulty sleeping 
causing inability to 
perform usual 
social & functional 
activities  
Disabling 
insomnia causing 
inability to 
perform basic 
self-care 
functions 
* Note that, while the grading scale provided for Unintentional Weight Loss may be used as a guideline when 
grading anorexia, this is not a requirement and should not be used as a substitute for clinical judgment.  
RESPIRATORY 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Bronchospasm 
(acute)  
FEV1 or peak flow 
reduced to 70 – 
80%  
FEV1 or peak flow 
50 – 69%  
FEV1 or peak flow 
25 – 49%  
Cyanosis OR FEV1 
or peak flow < 25% 
OR Intubation  
Dyspnea or 
respiratory 
distress  
Adult ≥ 14 
years  
Dyspnea on 
exertion with no or 
minimal 
interference with 
usual social & 
functional 
activities  
Dyspnea on 
exertion causing 
greater than 
minimal 
interference with 
usual social & 
functional 
activities  
Dyspnea at rest 
causing inability to 
perform usual 
social & functional 
activities  
Respiratory failure 
with ventilatory 
support indicated  


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
The contents of this document are confidential and proprietary to Quintiles Transnational Corp. Unauthorized use, disclosure or 
reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 88 of 91 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Dyspnea or 
respiratory 
distress  
Pediatric < 14 
years  
Wheezing OR 
minimal increase 
in respiratory rate 
for age  
Nasal flaring OR 
Intercostal 
retractions OR 
Pulse oximetry 90 
– 95%  
Dyspnea at rest 
causing inability to 
perform usual 
social & functional 
activities OR Pulse 
oximetry < 90%  
Respiratory failure 
with ventilatory 
support indicated  
MUSCULOSKELETAL 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Arthralgia  
See also 
Arthritis  
Joint pain causing 
no or minimal 
interference with 
usual social & 
functional activities  
Joint pain causing 
greater than 
minimal 
interference with 
usual social & 
functional activities  
Joint pain causing 
inability to perform 
usual social & 
functional activities  
Disabling joint 
pain causing 
inability to 
perform basic 
self-care 
functions  
Arthritis  
See also 
Arthralgia  
Stiffness or joint 
swelling causing no 
or minimal 
interference with 
usual social & 
functional activities  
Stiffness or joint 
swelling causing 
greater than 
minimal 
interference with 
usual social & 
functional activities  
Stiffness or joint 
swelling causing 
inability to perform 
usual social & 
functional activities  
Disabling joint 
stiffness or 
swelling causing 
inability to 
perform basic 
self-care 
functions  
Myalgia 
(non-
injection 
site)  
Muscle pain 
causing no or 
minimal 
interference with 
usual social & 
functional activities  
Muscle pain 
causing greater 
than minimal 
interference with 
usual social & 
functional activities  
Muscle pain 
causing inability to 
perform usual 
social & functional 
activities  
Disabling muscle 
pain causing 
inability to 
perform basic 
self-care 
functions  
 
 
 


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
The contents of this document are confidential and proprietary to Quintiles Transnational Corp. Unauthorized use, disclosure or 
reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 89 of 91 
LABORATORY: HEMATOLOGY 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Hemoglobin 
(Hgb)* 
10.0 – 10.9 g/dL 
6.18 – 6.79 
mmol/L  
OR Any decrease 
2.5 – 3.4 g/dL 1.58 
– 2.13 mmol/L  
9.0 – 9.9 g/dL 5.55 
- 6.17 mmol/L  
OR Any decrease 
3.5 – 4.4 g/dL 2.14 
– 2.78 mmol/L  
7.0 – 8.9 g/dL 4.34 
- 5.54 mmol/L  
OR Any decrease ≥ 
4.5 g/dL ≥ 2.79 
mmol/L  
< 7.0 g/dL < 4.34 
mmol/L  
Platelets, 
decreased  
100,000 – 
124,999/mm3 
100.000 x 109 – 
124.999 x 109/L  
50,000 – 
99,999/mm3 
50.000 x 109 – 
99.999 x 109/L  
25,000 – 
49,999/mm3 
25.000 x 109 – 
49.999 x 109/L  
< 25,000/mm3 < 
25.000 x 109/L  
WBC, 
decreased  
2,000 – 
2,500/mm3 2.000 
x 109 – 2.500 x 
109/L  
1,500 – 
1,999/mm3 1.500 
x 109 – 1.999 x 
109/L  
1,000 – 
1,499/mm3 1.000 
x 109 – 1.499 x 
109/L  
< 1,000/mm3 < 
1.000 x 109/L  
WBC, 
increased  
10,800 – 
15,000/mm3  
10.8 – 15.0 x 109/L  
15,001 – 
20,000/mm3  
15.1 – 20.0 x 109/L  
20,001 – 
25,000/mm3  
20.1 – 25.0 x 109/L  
>25,000/mm3  
>25.0 x 109/L  
*The decrease is a decrease from Baseline  
LABORATORY: CHEMISTRY 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Alkaline 
Phosphatase  
1.25 – 2.5 x 
ULN†  
2.6 – 5.0 x ULN†  
5.1 – 10.0 x 
ULN†  
> 10.0 x ULN†  
ALT (SGPT)  
1.25 – 2.5 x 
ULN  
2.6 – 5.0 x ULN  
5.1 – 10.0 x 
ULN  
> 10.0 x ULN  
AST (SGOT)  
1.25 – 2.5 x 
ULN  
2.6 – 5.0 x ULN  
5.1 – 10.0 x 
ULN  
> 10.0 x ULN  
Blood urea 
nitrogen 
(BUN)  
23 – 26 mg/dL  
8.3 – 9.5 
mmol/L  
27 – 31 mg/dL  
9.6 – 11.2 
mmol/L  
>31 mg/dL  
>11.2 mmol/L  
Requires dialysis  
Creatinine – 
elevated  
1.5 – 1.7 mg/dL  
121 - 145 
umol/L  
1.8 – 2.0 mg/dL  
146 – 170 
umol/L  
2.1 – 2.5 mg/dL  
171 – 208 
umol/L  
>2.5 mg/dL or requires 
dialysis  
>208 umol/L or requires 
dialysis  
Bilirubin 
(Total)  
1.1 – 1.5 x ULN  
1.6 – 2.5 x ULN  
2.6 – 5.0 x ULN  
> 5.0 x ULN  


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
The contents of this document are confidential and proprietary to Quintiles Transnational Corp. Unauthorized use, disclosure or 
reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 90 of 91 
LABORATORY: URINALYSIS 
Parameter 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Hematuria 
(microscopic)  
5 – 10 
cells/mm3  
 
+   
> 10 cells/mm3  
 
 
++/+++ 
Gross, with or 
without clots OR with 
RBC casts   
++/+++ 
Transfusion 
indicated  
 
++/+++ 
Proteinuria, 
random collection  
1 +  
2 – 3 +  
4 +  
NA  
Proteinuria, 24 
hour collection 
≥ 10 years  
200 – 999 
mg/24 h 
0.200 – 0.999 
g/d  
1000 – 1999 
mg/24 h 
1.000 – 1.999 g/d  
2000 – 3500 mg/24 h  
2.000 – 3.500 g/d  
> 3500 
mg/24 h  
> 3.500 g/d 
 
 
 


 
Document: 
Aeras_C-040-404_SAP_V2_0 
Author: 
Louise van Aswegen 
Version Number: 
2.0 
 
 
Version Date: 
29SEP2017 
Template No:  CS_TP_BS016 – Revision 3 
Reference:  CS_WI_BS005 
Effective Date: 01May2012 
 
Copyright © 2012 Quintiles Transnational Corp. All rights reserved. 
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reproduction is strictly prohibited. 
 
STATISTICAL ANALYSIS PLAN: PROTOCOL C-040-404 
Page 91 of 91 
APPENDIX 8. PROTOCOL-DEFINED VISIT WINDOWS 
Visit 
Visit Window 
Day 84 
84 ± 7 days from Day 0 
Month 6 
168 ± 14 days from Day 0 
Month 8 
168 ± 14 days from Day 56 
Month 9/12 
84/168 ± 14 days from initial conversion at Month 6 
Month 12 
336 ± 14 days from Day 0 
Month 15/18 
84/168 ± 14 days from initial conversion at Month 12 
Month 18 
504 ± 14 days from Day 0 
Month 21/24 
84/168 ± 14 days from initial conversion at Month 18  
Month 24 
672 ± 14 days from Day 0 
Month 27/30 
84/168 ± 14 days from initial conversion at Month 30