Clinical trial NCT02374814
Conditions: Rabies
Sponsor: State University of New York - Upstate Medical University
Start date: 2015-03-24; Completion date: 2016-09-22
Enrollment: 60 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
The purpose of this study is to compare the effectiveness of a two dose versus a three dose schedule and intramuscular versus intradermal injection for pre-exposure prophylaxis.

Detailed description:
This is an exploratory vaccine trial to evaluate immunogenicity of a non-licensed dosing schedule and route of administration for a currently FDA licensed rabies vaccine for pre-exposure prophylaxis against rabies infection. The goal of this study is to characterize the immune response and persistence of immunity to a shortened dose schedule and intradermal (ID) administration, relative to the current licensed dosing schedule of the rabies vaccine (3 dose (0, 7, 21 days) IM). Rabies virus is endemic throughout the world due to high rates of both wild and domestic animal rabies and the risk to deployed military in endemic areas is considerable. Currently the commonly supported pre-exposure prophylaxis regimen for rabies, in the United States is comprised of three, 1.0 ml intramuscular (IM) injections of the human diploid cell vaccine (HDCV) or purified chick embryo cell (PCEC) rabies vaccine on days 0, 7, and 21 or 28. Modified, two and three dose schedules of intradermal (ID) injections of 0.1 ml of HDCV and PCEC are utilized outside the US. These two and three dose intradermal schedules share a similar safety and immunogenicity profile to intramuscular vaccinations and are easily boosted at one year after vaccination. A death, from rabies, of a US Soldier returned from Afghanistan underscores the importance of rabies pre-exposure prophylaxis for soldiers and the need to evaluate the safest, most effective means of vaccinating large deploying forces. While the current three dose, 1 ml IM rabies series is effective, a shortened, equally effective vaccination series with significantly smaller dose per injection would greatly improve the logistics and cost associated with universal or even targeted coverage of deploying soldiers. Evaluation of a shorter, smaller-dose, pre-exposure vaccination series for rabies is the goal of this study.

Arms: arm 1: Intramuscular injection: 1mL at 0, 7 and 21 days. An additional 1 mL intramuscular dose at day 365 arm 2: Intradermal injection: 0.1mL at 0, 7 and 21 days. A single intramuscular 1 mL dose at day 365 arm 3: Intramuscular injection: 1mL at 0, 7 days. An additional 1 mL intramuscular dose at day 365 arm 4: Intradermal injection: 0.1mL at 0, 7 days. A single intramuscular 1 mL dose at day 365 arm 5: Albumin and saline comparator, Intramuscular injection: 1mL arm 6: Albumin and saline comparator, Intradermal injection: 0.1mL
Interventions: intervention 1: Rabies vaccine intervention 2: Placebo
Intervention descriptions: intervention 1: Compare dose schedule and route of administration intervention 2: Placebo

Protocol documents:
ICF_002.pdf: 
Title: Immunogenicity of a Two vs Three dose, Intradermal (ID) vs Intramuscular (IM) 
Administration of a Licensed Rabies Vaccine for Pre-Exposure Vaccination 
 
NCT number: 02374814 
 
 
Document date: 26 March 2015 


 
 
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SUNY Upstate Medical University  
 
Title of Study: Immunogenicity of a Two vs Three dose, Intradermal (ID) vs Intramuscular 
(IM) Administration of a Licensed Rabies Vaccine for Pre-Exposure Vaccination 
 
 
Consent / Authorization Form 
 
Background/Purpose: 
 
You are being asked to participate in a research study to help evaluate an investigational dosing 
schedule and route of administration for a Rabies vaccine, called RabAvert®, because you are a 
healthy individual between 18 and 60 years of age. This research study is being funded by the 
US Department of Defense, Walter Reed Army Institute for Research. 
 
Research studies include only those subjects who choose to take part. Please take your time to 
make your decision. Please ask the study doctor or the study staff to explain any words or 
information that you do not understand. You may also want to discuss it with your friends and 
family. 
 
The RabAvert® rabies vaccine is approved by the US Food and Drug Administration (FDA). 
The standard (approved) way to give this vaccine for the prevention of rabies is 3 doses given as 
an injection into the muscle.  In this study we will use the FDA approved vaccine; however, we 
are testing a different dosing schedule and different route of administration; which is considered 
investigational.    
 
We will compare two things: 
 
1) A two dose schedule (investigational) versus a three dose schedule (standard) of vaccine. 
2) Injection of the vaccine just under the skin (investigational) compared to injection of the 
vaccine into a muscle (standard). 
 
We plan to enroll 60 subjects in the study.  Subjects will be randomly assigned by chance to one 
of the six study groups shown below.  You will be told to which group you have been assigned.  
 


 
 
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Vaccine 
Group 
 
Day 
Number of subjects per 
Group 
0 
7 
  21 
365 
1 
IM 
IM 
 
IM 
IM 
12 
2 
ID 
ID 
 
ID 
IM 
12 
3 
IM 
IM 
 
IM 
12 
4 
ID 
ID 
 
IM 
12 
5 (control) 
IM 
 
 
 
6 
6 (control) 
ID 
 
 
 
6 
 
IM = intramuscular 
ID = Intradermal 
 
Study Procedures: 
 
The study will last for 13 months and will include 3-12 visits to the Clinical Research Unit 
(located on the Upstate campus) depending on what group you are in. 
 
At the first visit you will be evaluated by the study team to see if you are eligible for the study.  
This is called the screening visit and will include a medical evaluation. To be eligible you must: 
• 
Be at least 18 years old and no more than 60 years old 
• 
Be in good health, as determined by medical history, and physical examination. 
• 
If you are a female of child-bearing potential, you must have a negative urine pregnancy 
test result and be willing to use oral, implantable, transdermal or injectable contraceptives 
or another reliable method of contraception, approved by the study doctor, for the 
duration of the study and for 30 days after your last vaccination. 
- 
The study doctor or study staff will tell you if the pregnancy test results are 
positive.  
- 
The results of pregnancy testing must be negative in order for you to be in the 
study 
• 
Be able to give your informed consent  
• 
Come to the study center for all visits with the study doctor 
 
If you are eligible and want to continue to participate in the study, you will be randomly assigned 
to one of the six study groups. Your chance of being assigned to any group is two times higher 
for groups 1 to 4 then groups 5 or 6.  The six study groups are: 
• Group 1 which will receive four doses of vaccine on day 0, day 7, 21 and day 365. The 
vaccine will be administered intramuscularly (into the muscle) by needle and syringe.  
There will be 12 subjects in this group. 
• Group 2 which will receive four doses of vaccine on day 0, day 7, 21 and day 365. The 
vaccine will be administered intradermaly (under the skin) by needle and syringe on days 


 
 
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0, 7 and 21. The dose on day 365 will be administered intramuscularly by needle and 
syringe.  There will be 12 subjects in this group. 
• Group 3 will receive three doses of vaccine, on day 0, 7 and day 365. They will be 
administered intramuscularly by needle and syringe.  There will be 12 subjects in this 
group. 
• Group 4 will receive three doses of vaccine on day 0, 7 and day 365. They will be 
administered intradermaly by needle and syringe on days 0 and 7. The dose on day 365 
will be administered intramuscularly by needle and syringe.   There will be 12 subjects in 
this group. 
• Group 5 will receive one dose of human serum albumin on day 0. It will be administered 
intramuscularly by needle and syringe on day 0. There will be 6 subjects in this group. 
• Group 6 will receive one dose of human serum albumin on day 0. It will be administered 
intradermaly by needle and syringe on day 0. There will be 6 subjects in this group. 
 
You will need to stay at the study site for about 30 minutes after vaccination; these visits will be 
about one hour. 
 
You will be asked to come back at regular intervals for follow up visits and blood draws.  Most 
of these follow up visits will take 30 minutes. You will be provided with a schedule of the 
remaining visits at your Day 0 vaccination.  
 
Follow up blood draws will be between 3ml and 98 mls of blood ( max amount: about 6 1/2 
tablespoons). 
 
 
A further description of the tests/procedures that will be done while in the study (after screening) 
is explained below: 
 
• Physical examination based on your signs and symptoms of any illness, 
• Vital signs (temperature, heart rate, pulse and respiratory rate), 
• Urine test for pregnancy at each vaccination visit (females of child bearing potential 
only),  
• Blood test for presence of vaccine components, 
• Blood test for the presence of innate immune responses.  Innate immune responses are 
the first defense to fight infections caused by pathogens.  Innate immune responses 
provide an immediate but non-specific response. Innate immunity occurs naturally as a 
result of a person’s genetic constitution or physiology, and does not require previous 
exposure to the pathogen or vaccination, 
• Blood test for the presence of cellular immune responses.   
 


 
 
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• Cell-mediated immunity is an immune response that does not involve antibodies but 
rather involves the activation of specific cells in response to a foreign antigen.  These 
cells will recognize and kill infected cells, thereby providing protection from spread of 
infection, 
• Assessment of any adverse experiences (side effects) and changes in medications you are 
taking. 
 
The total volume of blood drawn for the entire study will be approximately 9-604 mls, depending 
on your assigned group.  For comparison, when people volunteer to donate blood, the volume is 
about 250-450 ml at one time.  Most of your samples will be used for tests described above. We 
will also be looking at how differences in each person’s immune system may affect their 
response to the vaccine.  To do this we will be looking at your HLA type.  HLA means human 
leukocyte antigen.  Your HLA type is defined by a group of genes that are important elements 
for immune function.  This is considered a type of genetic testing.   
 
While this testing will reveal genetic information about you, the only genetic information we are 
looking for is the genetic information that affects your immune response to the vaccine.  You 
may have heard of genetic testing that is used in determining biologic parents or a person’s 
ability to get or keep medical and/or life insurance.  We will not be collecting or storing this type 
of genetic information and since the significance of these tests is not known for you, we will not 
release the results of any genetic testing.   There will be no other genetic testing done, other than 
HLA typing. 
 
Saving Leftover Samples for Future Research (OPTIONAL): 
 
Any blood sample left over may be used for future research studies, such as additional analysis. 
These samples will be stored (possibly indefinitely) at a designated facility and provided to 
scientists performing research.  Your stored sample will be coded with a unique number for this 
study and your identity will only be known by the study site.  Saving these leftover blood 
samples for future research is OPTIONAL and you may choose to allow us to save your samples 
or not to save your samples. Your decision about whether or not to allow your samples to be 
saved for future research will have no affect on your participation in the rest of the study.  No 
genetic testing of any kind will be done on your leftover samples. 
Please let us know whether you allow us to save your samples to perform such studies by 
responding the following: 
 
 I agree to allow my blood samples collected in this study to be saved for future related 
research. 
 
 I do not agree to allow my blood samples collected in this study to be saved for future 
related research. 
 
 
 


 
 
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Are there things you should not do while participating in the study? 
There are some restrictions if you decide to participate in this study.  You should not receive any 
other vaccines in the 4 weeks before the first vaccination is given and in the 4 weeks after the 
last vaccination.  If you participate in this study, you should not be involved in other vaccine 
studies or other clinical studies (involving drugs or other agents, or medical devices) until one 
month after the last visit of this study.  
  
Risks/Discomforts: 
 
Overall, the vaccine is well tolerated. Reactions to the current rabies vaccines are mild with local 
pain, redness, and swelling at the injection site.  Some general reactions occurring less frequently 
are headache and fatigue.  
Rabies vaccine is the first vaccine to have been developed by Louis Pasteur as a rabbit spinal 
cord vaccine.  The rabies vaccine is currently available in the U.S. as a human diploid cell 
vaccine (HDCV) or as a purified chick embryo cell vaccine (PCECV).  The vaccine being used 
for this study, RabAvert® is a PCECV, and is used for pre and post exposure. It is made by 
Novartis Pharmaceuticals.  
Human serum albumin is a liquid diluent that has a similar composition of your body’s blood 
make-up. This diluent is used as a form of blood component replacement for people in the 
hospital. It is made by Greer Labs, Inc. 
 
Could you have an allergic reaction? 
Sometimes people have allergic reactions to vaccines.  If you have a very bad allergic reaction, 
you could die.  Some things that happen during an allergic reaction that could be a sign or 
symptom of a life-threatening allergic reaction (anaphylaxis) are: 
• a rash  
• having a hard time breathing 
• wheezing  
• a sudden drop in blood pressure (making you feel dizzy or lightheaded) 
• swelling around the mouth, throat, or eyes 
• a fast pulse 
• sweating 
You should get medical help right away and contact the study doctor or study staff if you have 
any of these or any other side effects during the study. 
For female participants of child bearing potential: 
All women of child bearing potential participating in the study will be asked to have a pregnancy 
test before each vaccination. You should also not participate in this study if you are nursing a 
child.  The effects of the vaccine on nursing children are not known at this time. 
Any woman who finds that she has become pregnant while taking part in the study should 
immediately inform the study doctor.   


 
 
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The person will not receive further vaccinations and they will be followed-up to determine the 
outcome of the pregnancy.  
 
There are some known risks involved with participating in this study: 
 
• Vaccinations:  
• Subjects may have discomfort associated with intradermal or intramuscular 
injection of the vaccine.  The injections will be in the forearm for ID and in the 
deltoid for IM.  
• Subjects may develop a flu-like illness with fever and body aches after 
vaccination.  This is similar to other vaccines for viruses. 
 
• Blood draws: Subjects may have pain and/or bruising at the location on your arm where 
the blood was taken. On rare occasions, it may cause lightheadedness or fainting or 
infection. 
 
 
Benefits: 
 
You may or may not benefit from participating in this study. We do not know if giving 
the rabies vaccine in this way will allow your body to develop protective antibodies to the 
rabies virus. Participation in this study does not mean you are protected against Rabies 
virus and any possible exposure to Rabies virus should be treated with the standard (FDA 
approved) post exposure treatment regimen. The information obtained from this study 
may help confirm a new effective dosing and route administration of the rabies vaccine to 
help protect a greater population in the future. 
 
Voluntary Participation/ Study Withdrawal: 
 
Your participation in this study is entirely voluntary and you may refuse to participate or 
withdraw from the study at any time, without penalty or loss of benefits to which you would 
normally be entitled. Your decision about whether or not to participate in the study will not 
affect your relationship with SUNY Upstate Medical University.  
 
The Study Doctor can take you off the study without your consent if: 
• 
He or she feels it is for your benefit and/or safety, for example if a side effect or 
medical condition develops during the study.   
• 
You are unable to follow the study procedures. 
• 
It is discovered that you do not meet the study requirements. 
• 
If the study is stopped by the Study Doctors or other oversight committees before 
the end of the study 
If there are significant new findings or information that might change your mind about 
participating in the study, we will give you the information and allow you to reconsider whether 
or not to continue. 


 
 
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If you decide to stop taking part in this study, you should tell the study doctor.  If you stop being 
in the study early, the study doctor or study staff may ask you some questions about being in the 
study.  To help you leave the study safely, the study doctor may ask you to participate in more 
tests. 
If the study is stopped early or if you choose to stop participating in the study, you are 
encouraged to return to the study site for these assessments: 
- 
Physical examination 
- 
Injection site assessment (if you leave the study within 14 days of vaccination) 
- 
Blood test for presence of study vaccine components (if you leave the study within 14 
days of study vaccination) 
  
 
Alternatives: 
 
You can receive the standard rabies vaccine outside this research study.  If you feel you should 
be treated with the rabies vaccine, you should discuss this with your regular doctor. If you decide 
not to participate in this research study, you may still enroll in other research studies. 
 
Costs/Payments: 
 
There are no costs to you or your insurance carrier for participating in this study. All costs for the 
required study visits, examinations, laboratory procedures and study vaccine will be paid by the 
study.  
 
You will be paid for each visit to cover your expenses. You will be paid $50 per visit for every 
visit you complete beyond the screening visit. Total payment for the study will be between $150 
and $600 dollars depending on which vaccination group you are assigned.  In the event that your 
participation in the study is discontinued early, you will only be paid for the visits completed.  
 
In addition, by accepting payment for participating in this study, certain identifying information 
about you may be made available to professional auditors to satisfy audit and Federal reporting 
requirements, but confidentiality will be preserved. Please note that if you earn $600 or over in a 
calendar year as a research subject, you may have to pay taxes on these earnings. 
 
 
Questions: 
 
If you have any questions about the research, or in the event of a research-related injury, please 
contact Dr. Mark E. Polhemus at (315) 464-9493. If you have any questions about your rights as 
a research subject, please contact the SUNY Upstate Medical University Institutional Review 
Board Office at (315) 464-4317. 
 
 
 


 
 
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In Case Of Injury: 
 
In the event of illness or physical injury resulting from taking part in this research study, medical 
treatment will be provided at University Hospital. You will be responsible for any costs not paid 
by your insurance company. No other compensation is offered by SUNY Upstate Medical 
University. SUNY Upstate Medical University has no plans to give you money if you are 
injured. You have not waived any of your legal rights by signing this form. 
 
 
 
Confidentiality of Records and Authorization to Use/Share Protected Health Information 
for Research: 
 
If you agree to participate in this research, identifiable health information about you will be used 
and shared with others involved in this research. For you to be in this research we need your 
permission to collect and share this information. Federal law protects your right to privacy 
concerning this information.  
 
When you sign this consent form at the end, it means that you have read this section and 
authorize the use and/or sharing of your protected health information as explained below. Your 
signature also means you have received a copy of Upstate’s Notice of Privacy Practices.  
 
Individually identifiable health information under the federal privacy law is considered to be any 
information from your medical record, or obtained from this study, that can be associated with 
you, and relates to your past, present, or future physical or mental health or condition. This is 
referred to as protected health information. 
 
Your protected health information will be kept confidential. Your identity will not be revealed in 
any publication or presentation of the results of this research. 
 
A description of this clinical trial will be available on http://www.clinicaltrials.gov, as required 
by U.S. Law. This Web site will not include information that can identify you. At most, the Web 
site will include a summary of the results. You can search this Web site at any time. 
 
 
Why is it necessary to use/share your protected health information with others? 
 
The main reason to use and share your health information is to conduct the research as described 
in this consent form. Your information may also be shared with people and organizations that 
make sure the research is being done correctly, and to report unexpected or bad side effects you 
may have. 
 
In addition, we may be required by law to release protected health information about you; for 
example, if a judge requires such release in a lawsuit, or if you tell us of your intent to harm 
yourself or others. 
 


 
 
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What protected health information about you will be used or shared with others as part of 
this research? 
 
We may use and share the results of tests, questionnaires, and interviews. We may also use and 
share information from your medical and research records. We will only collect information that 
is needed for the research. 
 
Who will be authorized to use and/or share your protected health information? 
 
The researchers, their staff and the staff of Upstate Medical University participating in the 
research will use your protected health information for this research study. In addition, the 
Upstate Institutional Review Board (IRB), a committee responsible for protecting the rights of 
research subjects, and other Upstate Medical University or University Hospital staff who 
supervise the way the research is done may have access to your protected health information.  
 
The researchers and their staff will determine if your protected health information will be used or 
shared with others outside of Upstate Medical University for purposes directly related to the 
conduct of the research.  
 
With whom would the protected health information be shared? 
 
Your protected health information may be shared with: 
• Representatives from the US Department of Defense, funders of this research; 
• Collaborators at the Walter Reed Army Institute of Research; 
• Kansas State University Rabies Laboratories;   
• Frontier Science, database management group 
• The research monitor; 
• Federal agencies that supervise the way the research is conducted, such as the 
Department of Health and Human Services’ Office for Human Research Protections, the 
Food and Drug Administration (FDA) or other governmental offices as required by law.   
 
All reasonable efforts will be used to protect the confidentiality of your protected health 
information. However, not all individuals or groups have to comply with the Federal privacy 
law. Therefore, once your protected health information is disclosed (leaves Upstate Medical 
University), the Federal privacy law may not protect it.   
 
For how long will your protected health information be used or shared with others? 
 
There is no scheduled date at which this information will be destroyed or no longer used. This is 
because information that is collected for research purposes continues to be used and analyzed for 
many years and it is not possible to determine when this will be complete. 
 
 


 
 
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Can you withdraw your authorization to collect/use/share your protected health 
information? 
 
You always have the right to withdraw your permission (revoke authorization) for us to use and 
share your health information, by putting your request in writing to the investigator in charge of 
the study.  This means that no further private health information will be collected. Once 
authorization is revoked, you may no longer participate in this research activity, but standard 
medical care and any other benefits to which you are entitled will not be affected. Revoking your 
authorization only affects uses and sharing of information obtained after your written request has 
been received, but not information obtained prior to that time.  
 
Even after you withdraw your permission, Upstate Medical University may continue to use and 
share information needed for the integrity of the study; for example, information about an 
unexpected or bad side effect you experienced related to the study.  
 
Can you have access to your health information? 
 
At the end of the study, you have the right to see and copy health information about you in 
accordance with the SUNY Upstate Medical University policies; however, your access may be 
limited while the study is in progress. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


 
 
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Consent To Participate In Research & Authorization To Use And Share Personal Health 
Information: 
 
I hereby give my consent to participate in this research study and agree that my personal health 
information can be collected, used and shared by the researchers and staff for the research study 
described in this form. I will receive a signed copy of this consent form. 
  
______________________________________ 
 
 
__________________   
Signature of subject  
 
 
 
 
 
Date 
 
_______________________________________ 
 
 
Printed name of subject 
 
 
 
 
 
  
 
___________________________________________ 
 
__________________ 
Signature of Person Obtaining Consent/Authorization 
 
Date 
 
 
_______________________________________ 
 
 
Name of Person Obtaining Consent/Authorization 
 
 
 

Prot_000.pdf: 
 
 
 
 
 
 
Title: Immunogenicity of a Two vs Three dose, Intradermal (ID) vs Intramuscular (IM) Administration 
of a Licensed Rabies Vaccine for Pre-Exposure Vaccination 
 
NCT number: 02374814 
 
Document date: 02 June 2016 
 
 
 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
 
Immunogenicity of a Two vs Three dose, Intradermal (ID) vs Intramuscular (IM) 
Administration of a Licensed Rabies Vaccine for Pre-Exposure Vaccination 
 
Randomized, open-label, single center trial in adults aged 18 to 60 years in the US 
 
 
 
 
Clinical Trial Protocol 
                                                      Version 6.0 dated 20 MAY 2016 
 
 
 
Funder’s Representative: 
LTC(P) Paul Keiser 
 
Walter Reed Army Institute of Research 
 
503 Robert Grant Avenue 
 
Silver spring, MD 20910 
 
 Principal Investigator: 
Mark E Polhemus 
 
Upstate Medical University 
 
725 Irving St. Suite 311 
 
Syracuse, NY 13210 
 
 
 
Licensed Products: 
 The RabAvert® Rabies Vaccine                                         
 
Form /Route: 
Intradermal and Intramuscular 
 
Indication For This Study: 
Comparison of Dose and Schedule  
 
                
 
Research Monitor: 
Tasaduq Fazili, MD 
 
Upstate Medical University  
                                                                     725 Irving Ave., Suite 311 
                                                                     Syracuse, NY 13210 
 
 
 
 
 
 
 
 
 
Funder’s Clinical Development 
           Lou Jasper 
Director: 
 USAMMDA 
 
 1430 Veteran’s Drive 
 
 Ft Detrick, MD 21702 
 
 Tel: (301) 619-4951 
 
 Fax: (301) 619-2304 
 
 
 
Study Site/PI Location: 
Center for Global Health and Translational Science  
  
Upstate Medical University 
 
Institute for Human Performance 
                                                              505 Irving Avenue 
  
Syracuse, NY 13210 
 
 
 
 
 
 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
 
                                            Table of Contents 
 
Synopsis………………………………………………………………………………………………….4 
List of Terms and Abbreviations……………………………………………………………………..23 
1   Introduction…………………………………………………………………………………...........25 
 
1.1 Background of Rabies……………………………………………………………………..25 
 
1.2 Background of the Licensed Product…………………………………………………….26 
 
1.3 Potential Benefits to Subjects……………………………………………………………..27 
 
1.4 Potential Risks to Subjects………………………………………………………………..27 
 
1.5 Rationale…………………………………………………………………………………...28 
2    Objectives………………………………………………………………………………………….29 
 
2.1 Primary Objectives………………………………………………………………………..29 
 
2.2 Secondary Objectives……………………………………………………………………..29 
 
2.3 Exploratory Objectives…………………………………………………………………...30 
3    Investigators and Trial Organization……………………………………………………………30 
4    Independent Ethics Committee/ Institutional Review Board…………………………………..30 
5    Trial Design………………………………………………………………………………………..30 
 
5.1 Justification of the Trial Design………………………………………………………….31 
 
5.2 Trial Plan…………………………………………………………………………………..32 
 
5.3 Visit Procedures…………………………………………………………………………...32 
 
5.4 Planned Trial Calendar…………………………………………………………………...45 
6    Enrollment and Retention of Trial Population………………………………………………….45 
 
6.1 Recruitment Procedures…………………………………………………………………..45 
 
6.2 Informed Consent Procedures……………………………………………………………45 
 
6.3 Screening Criteria…………………………………………………………………………45 
 
6.4 Inclusion Criteria………………………………………………………………………….46 
 
6.5 Exclusion Criteria…………………………………………………………………………46 
 
6.6 Medical History…………………………………………………………………………....47 
 
6.7 Contraindications for Subsequent Vaccinations………………………………………...47 
 
 
6.7.1 Temporary Contraindications………………………………………………….47 
 
 
6.7.2 Definitive Contraindications……………………………………………………47 
 
6.8 Other Conditions-for-Withdrawal beyond Definitive Contraindications……………..48 
 
6.9 Lost to Follow-up Procedures…………………………………………………………….49 
 
6.10 Classification of Subjects Who Discontinue the Trial…………………………………49 
 
 
6.10.1 Follow-up of Discontinuations………………………………………………...49 
 
 
6.10.2 Follow-up and Reporting of Pregnancies…………………………………….49 
 
6.11 Modification of the Trial and Protocol…………………………………………………50 
7       Safety Emergency Coverage……………………………………………………………………50 
8       Vaccines Administered………………………………………………………………………….50 
 
8.1 Identity of Trial Product: Rabies Vaccine……………………………………………….51 
 
 
8.1.1 Composition……………………………………………………………………...51 
 
 
8.1.2 Preparation and Administration………………………………………………..52 
 
 
8.1.3 Dose Selection and Timing………………………………………………………52 
 
 
8.1.4 Labeling and Packaging…………………………………………………………52 
 
 
8.1.5 Product Shipment………………………………………………………………..52 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
 
 
 
8.1.6 Product Storage…………………………………………………………………52 
8.1.7 Product Accountability…………………………………………………………52 
8.1.8 Replacement Doses……………………………………………………………...52 
8.1.9 Disposal of Unused Products…………………………………………………...52 
9        Blinding and Code-breaking Procedures……………………………………………………..53 
10      Randomization and Allocation Procedures…………………………………………………...53 
11      Treatment Compliance…………………………………………………………………………53 
12      Concomitant Medication or Other Therapies………………………………………………...53 
13      Management of Samples………………………………………………………………………..55 
 
13.1 Sample Collection………………………………………………………………………...55 
 
 
13.1.1 Blood Samples…………………………………………………………………..55 
 
 
13.1.2 Additional Biological Samples…………………………………………………55 
 
13.2 Sample Preparation………………………………………………………………………55 
 
13.3 Sample Storage and Shipment…………………………………………………………..56 
 
13.4 Future Use of Stored Serum Samples for Research……………………………………56 
14      Clinical Supplies………………………………………………………………………………...56 
15      Endpoints and Assessment Methods…………………………………………………………...56 
 
15.1 Primary Endpoints……………………………………………………………………….56 
 
 
15.1.1 Immunogenicity Endpoints……………………………………………………56 
 
15.2 Secondary Endpoints…………………………………………………………………….57 
 
 
15.2.1 Immunogenicity Endpoints……………………………………………………57 
 
15.3 Assessment Methods……………………………………………………………………..57 
16       Reporting AEs to IRBs………………………………………………………………………...57 
17       Reporting of Serious Adverse Events…………………………………………………………57 
           Timelines and process for SAE reporting…………………………………………………….57 
           Process for SAE documentation………………………………………………………………58 
           Timelines and process for pregnancy reporting……………………………………………..58 
 
17.1 Follow-up Reporting by the Investigator……………………………………………….58 
 
17.2 Reporting of SAEs Occuring After a Subject Has Completed the Study…………….58 
 
17.3 Assessment of Causality………………………………………………………………….58 
 
17.4 Reporting SAEs to Health Authorities and IRBs………………………………………59 
18       Data……………………………………………………………………………………………...59 
 
18.1 Data Collection and eCRF Completion…………………………………………………59 
 
18.2 Data Management………………………………………………………………………..60 
 
18.3 Data Review………………………………………………………………………………60 
19       Statistical Methods……………………………………………………………………………..60 
 
19.1 Statistical Methods for Primary Immunogenicity Objectives…………………………61 
 
19.2 Statistical Methods for Secondary Immunogenicity Objectives………………………61 
 
19.3 Per-Protocol Analysis Set………………………………………………………………..61 
 
19.4 Full Analysis Set………………………………………………………………………….62 
 
19.5 Populations Used in Analysis…………………………………………………………….62 
20      Handling of Missing Data and Outliers………………………………………………………..62 
 
20.1 Immunogenicity…………………………………………………………………………..62 
21      Determination of Sample Size and Power Calculation……………………………………….63 
22      Ethical and Legal Issues and Investigator/Sponsor Responsibilities………………………...63 
 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
 
22.1 Ethical Conduct of the Trial/Good Clinical Practice…………………………………..63 
 
22.2 Source Data and Source Documents…………………………………………………….63 
 
22.3 Confidentiality of Data and Access to Subject Records………………………………..63 
 
22.4 Monitoring, Auditing, and Archiving…………………………………………………...64 
 
 
22.4.1 Monitoring……………………………………………………………………...64 
 
 
22.4.2 Audits and Inspections………………………………………………………....64 
 
 
22.4.3 Archiving………………………………………………………………………..64 
23       Stipends for Participation……………………………………………………………………...65 
24       Reference List…………………………………………………………………………………..65 
 
 
 
  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
Synopsis 
 
Licensed Product: 
RabAvert® Rabies Vaccine 
 
Title of the Trial: 
Immunogenicity of a two vs three dose, intradermal (ID) vs intramuscular (IM) 
administration of rabies vaccine for pre-exposure vaccination 
Trial Centers: 
Single site trial conducted at the State University of New York (SUNY) Upstate Medical University 
Center for Global Health and Translational Science 
Planned Trial 
Period: 
February 2014: IRB Submission 
Planned trial period - FVFS (first visit, first subject) to LCLS (last contact, last subject): October 2014  
February 2016  
October2014: Enrollment 
October2014: First subject, first vaccination 
January  2015: Last subject, last vaccination 
February 2016: Last contact, last subject/Final study visit 
July 2016: Assays Complete 
January 2017: Final clinical study report submission 
 
Trial Design and 
Methodology: 
This will be a randomized, open-label, single center study involving60healthy adults aged ≥ 18 years 
to ≤ 60 years at the time of first vaccination. 
 
Table S1: Schedules of vaccine administration and number of subjects by group 
Vaccine 
Group 
Day 
 
Number per Group 
0 
7 
21 
  365 
(Boost) 
1 
IM 
IM 
IM 
IM 
12 
2 
ID 
ID 
ID 
IM 
12 
3 
IM 
IM 
 
IM 
12 
4 
ID 
ID 
 
IM 
12 
5 
IM 
control 
 
 
 
6 
6 
ID 
control 
 
 
 
6 
In all groups, subjects will provide blood samples for determination of neutralizing antibody (Ab) titers.  
See attached Tables of Study Procedures for summary of assays for each visit.  


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
 
Subjects will be randomly assigned to one of the six groups through a computer generated process. A 
standard permuted-block design will be used for randomization/allocation in order to avoid undesirable 
differences in the numbers of patients assigned to each group. The randomization schedule will be 
generated by our statistician using SAS 9.3 PROC PLAN. Variable block sizes will be kept confidential 
from the protocol to reduce the predictability of the treatment assignments.  
After screening visit, eligible subjects that are randomized will be assigned a unique subject ID. The 
subject ID will remain unchanged throughout the study and the nurse will confirm the number at every 
visit. No single subject will receive more than one subject ID. Blood will be drawn at each follow-up 
visit to be used in research assays for antibody response, and proteomics evaluation.  See blood draw 
table for timing and volumes of blood draws and assays for which they will be used.  
In all groups additional blood samples may be taken if required to assess AEs, SAEs, or abnormal 
laboratory findings.  
 
 
Primary 
Objective: 
 
Immunogenicity 
1) To describe the percentage of subjects achieving a protective humoral immune response at 12 
months across immunization routes at 7 days post boost.  
Primary 
Endpoints: 
Immunogenicity 
1) Percentage of subjects in each group with titer of IgG antibodies above the protective 
limits ( > or = 0.5 IU/ml) at 12 months, 7 days after final vaccine dose. 
 
Secondary 
Objectives: 
Immunogenicity 
1) To describe the percentage of subjects maintaining a protective humoral immune response at 
12 months across immunization routes.  
Secondary 
Endpoint: 
 
Immunogenicity 
1) 
Percentage of subjects in each group with titer of IgG antibodies above the protective limits (
> or = 0.5 IU/ml) at 12 months, prior to boost.  
 
 
Exploratory 
Objectives 
Immunogenicity 
1) Describe the cell-mediated immune response to rabies vaccination in groups 1-4. 
 
2) Proteomic characterization of the host response before and after each vaccination to identify 
biomarkers and correlates of immunity 
Exploratory 
Endpoints 
Immunogenicity 
1) Frequency, kinetics, and/or phenotype of virus immune cell subsets (e.g., B cells, T cells) at 
baseline and in response to rabies vaccination in groups 1-4. 
2) Development of Proteome 3D for complete seroconversion, incomplete seroconversion, and 
severe reaction. 
3) Additional analyses not specified here may be performed in order to further characterize 
vaccine responses. 
 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
 
Planned 
Sample Size: 
A total of 60 subjects will be vaccinated. 
There will be four groups (Groups 1, 2, 3, 4,with 12 subjects in each group) and two groups (Groups 5 and 
6 with 6 subjects per group.) 
Schedule of  
Study 
Procedures: 
Screening visit: 
 All screened subjects will review and sign an informed consent form, and undergo a thorough medical 
history and physical examination to assess for inclusion/exclusion criteria. All females will provide a urine 
sample to assess for pregnancy.  Screening will be conducted within 60 days of vaccination.  Screening 
numbers will be assigned when the subject signs the consent document starting with screening #100, 101, 
102, etc.  When the subject has completed the screening visit and is eligible and willing to continue 
participation, they will be randomized according to the randomization process outlined in section 6.4.   
Screen-fails will not be recorded in the data management system for the study.    
 Vaccinations and Blood Sampling: 
All subjects will provide a blood sample before the first vaccination for baseline research labs.  Results of 
vaccination day labs will not be reviewed before immunization. 
Group 
Schedule of Vaccination and Blood Draw 
       
1     
 
 
Vaccinations: Rabies vaccine IM 3 doses. 
 
Blood samples:  Blood samples will be taken for research assays evaluating, humoral 
immunity, cell mediated immunity, and proteomics. See Blood Draw Table: Group 1 for timing 
of blood draws. 
 
Total blood volume for Group 1: approx 593 mL 
2 
 
Vaccinations: Rabies vaccine ID 3 doses  
 
Blood samples:  Blood samples will be taken for research assays evaluating humoral 
immunity, cell mediated immunity, and proteomics.  See Blood Draw Table: Group 2 for 
timing of blood draws. 
 
Total blood volume for Group 2: approx 593 mL 
3 
 
Vaccinations: Rabies vaccine IM 2 doses  
 
Blood samples:  Blood samples will be taken for research assays evaluating humoral 
immunity, cell mediated immunity, and proteomics. See Blood Draw Table: Group 3 for timing 
of blood draws. 
 
Total blood volume for Group 3: approx 510 mL 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
4 
 Vaccinations: Rabies vaccine ID 2 doses  
Blood samples:  Blood samples will be taken for research assays evaluating humoral 
immunity, cell mediated immunity, and proteomics.  See Blood Draw Table: Group 4 for 
timing of blood draws. 
 
Total blood volume for Group 4: approx 510 mL 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Vaccination: Placebo IM 1 dose 
Blood Samples: Blood samples will be taken for research assays evaluating proteomics.  
See Blood Draw Table: Group 5 for timing of blood draws.  
Total blood volume for Group 5: approx 9 mL 
5 
Vaccination: Placebo IM 1 dose 
Blood Samples: Blood samples will be taken for research assays evaluating proteomics. 
See Blood Draw Table: Group 6 for timing of blood draws.  
Total blood volume for Group 6: approx 9 mL 
6 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
 
Duration of the 
Trial: 
The expected duration of a subject’s participation in the trial will be 13 months for subjects in Groups 
1, 2, 3, and 4. In groups 5 and 6 participation will end after Day 7.  
Licensed Product:
 
Form: 
 
 
 
 
Composition: 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Route: 
 
 
 
Placebo: 
 
 
Form: 
 
Composition: 
 
Route: 
RabAvert® Rabies Vaccine 
RabAvert®: ≥2.5 units [contains albumin (human), amphotericin B (may have trace amounts), bovine 
gelatin, chicken egg protein, chlortetracycline (may have trace amounts), neomycin (may have trace
amounts); PCEC; grown in chicken fibroblast culture; supplied with diluent]. 
RabAvert® (Rabies Vaccine) produced by Novartis Vaccines and Diagnostics GmbH is a sterile 
freeze-dried vaccine obtained by growing the fixed-virus strain Flury LEP in primary cultures of 
chicken fibroblasts.  The strain Flury LEP was obtained from American Type Culture Collection as the 
59th egg passage.  The growth medium for propagation of the virus is a synthetic cell culture medium 
with the addition of human albumin, polygeline (processed bovine gelatin) and antibiotics.    The virus 
is inactivated with β-propiolactone, and further processed by zonal centrifugation in a sucrose density-
gradient.  The vaccine is lyophilized after addition of a stabilizer solution which consists of buffered 
polygeline and potassium glutamate.    One dose of reconstituted vaccine contains less than 12 mg 
polygeline (processed bovine gelatin), less than 0.3 mg human serum albumin, 1 mg potassium 
glutamate and 0.3 mg sodium EDTA.  Small quantities of bovine serum are used in the cell culture 
process.  Minimal amounts of chicken protein may be present in the final product; ovalbumin content is
less than 3 ng/dose (1 mL), based on ELISA.  In the final vaccine, neomycin is present at < 1 μg, 
chlortetracycline at < 20 ng, and amphotericin B at < 2 ng per dose.    RabAvert® is intended for 
intramuscular (IM) injection.  The vaccine contains no preservative and should be used immediately 
after reconstitution with the supplied Sterile Diluent for RabAvert® (Water for Injection).    The 
potency of the final product is determined by the US National Institute of Health (NIH) mouse potency 
test using the US reference standard. 
After reconstitution the vaccine should be administered immediately. The potency of 1.0 ml of 
RabAvert Rabies vaccine is equal to or greater than 2.5 international units of rabies antigen.  The 
Intramuscular (IM) dose is 1.0 ml and the intradermal (ID) dose is 0.1 ml.  
Human Serum Albumin (HSA) Saline 
Normal Saline with 0.03%. human serum albumin added. 
Produced by Greer Labs, for use as a sterile diluent.  
The Intramuscular (IM) dose is 1.0 ml and the Intradermal (ID) dose is 0.1 ml. 
 
Inclusion 
Criteria: 
 
Subjects must fulfill all of the following criteria in order to be eligible for trial enrollment: 
1)  Male and non-pregnant females aged ≥ 18 to ≤ 60 years on the day of inclusion 
2)  Able to comprehend and give informed consent  
3) Able to attend all scheduled visits and to comply with all trial procedures   
4)  Subject in good health, based on medical history and physical examination 
 
 


Rabies Vaccine   
RABVAX Version 6.0 20 MAY 2016 
 
Exclusion 
Criteria 
A subject fulfilling any of the following criteria is to be excluded from trial enrollment: 
 
1. Subject is pregnant, or lactating, or of childbearing potential (to be considered of non-childbearing 
potential, a female must be post- menopausal for at least 1 year, surgically sterile, or using an 
effective method of contraception or abstinence from at least 4 weeks prior to the first vaccination 
and until at least 4 weeks after the last vaccination).   
2. Participation in the 4 weeks preceding the first trial vaccination, or planned participation during 
the present trial period, in another clinical trial investigating a vaccine, drug, medical device, or 
medical procedure. 
3. Previous history of receiving the rabies vaccine. 
4. Previous history of receiving rabies immune globulin. 
5. Any major psychiatric disorder, such as severe depression, severe anxiety disorder, psychosis, 
schizophrenia, other major psychiatric disorders, or seizures. History of mild depression or anxiety 
disorder that is well controlled is not an exclusion criteria. 
6. Any history of cardiac arrhythmias, such as: Bradycardia, tachycardia, heart block, SVT, PAC, 
VF, VT, or any other conduction abnormalities. 
7.  Use of any immunosuppressive drug , including topical steroids of potency groups I, II or III 
within 30 days of the study period. 
8. Any immunosuppressive disorder, such as HIV, common variable, active cancers or 
chemotherapy. 
9. History of renal insufficiency or requiring dialysis. 
10. Any condition that would, in the opinion of the site investigator, place the subject at an 
unacceptable risk of injury or render the subject unable to meet the requirements of the protocol. 
11. Identified as an employee of the Investigator or study center, with direct involvement in the 
proposed study or other studies under the direction of that Investigator or study center, as well as 
family members (i.e., immediate, husband, wife and their children, adopted or natural) of the 
employee or the Investigator. 
 
Temporary Exclusion Criteria: Moderate or severe acute illness/infection (according to investigator 
judgment) on the day of vaccination or febrile illness (temperature ≥ 38.0°C [≥ 100.4°F]). A 
prospective subject should not be included in the study until the condition has resolved or the febrile 
event has subsided.  If the delay for the febrile illness exceeds the window between screening and 
vaccination, or if deemed necessary by the investigator, a prospective subject may be re-screened 
once the fever has resolved. 
 
 
 


Confidential/Proprietary Information 
Rabies Vaccine  
RABVAX Version 6.0 20 MAY 2016 
  
Page 12 of 69 
 
 
Statistical 
Methods:  
Immunogenicity –Secondary objectives: 
 
Immunogenicity will be assessed descriptively using the following parameters: 
 
Time points: Baseline, 28 days and 365 days after initial vaccine dose 
 Percentage of subjects in each group with titer of IgG antibodies above the protective limits (≥ 
0.5 IU/ml) at 365 days, prior to boost. 
 
 
 
 
 
Statistical 
Methods: 
This will be a descriptive study to assess immunogenicity of a licensed rabies vaccine using test dosage 
schedules and routes of administration. No formal statistical hypothesis tests will be conducted. 
Descriptive analyses will be based on the per-protocol analysis sets. 
Exploratory statistical analyses of final data may be conducted, if indicated by the descriptive results. 
Parametric, non-parametric and resampling (bootstrapping) methods for statistical inference may be used 
in exploratory analyses, based on data compliance with assumptions of methods.  P -values  ≤ 0.05 will be 
considered significant and p-values ≤ 0.10 will be considered a trend. Confidence intervals will be 
constructed at α=0.05 and α=0.10.  When necessary, p-value corrections for multiple comparisons will be 
applied. 
Descriptive and any inferential analyses will be carried out using SAS Version 9.2 (or more recent 
versions), which is licensed and supplied by SAS Institute, Cary, NC, USA. 
 
Immunogenicity – Primary objectives:  
 
Immunogenicity against rabies virus will be assessed descriptively using the following 
parameters: 
Timepoints:  Baseline, 28 days after initial dose of rabies vaccine and 7 days after a booster dose 
given at one year (372 days) 
 
• 
Number and percentage of subjects with a titer ≥ 0.5 IU/ml against rabies virus. 
 
 
Statistical 
Methods: 
Calculation of Sample Size: 
 
  The planned sample size is 60 subjects, who will be randomly allocated (2:2:2:2:1:1)to four treatment 
groups and two control groups (i.e.  12 subjects in groups 1, 2, 3, and 4 and 6 subjects in groups 5 and 
6) The sample size was chosen empirically, based on sample sizes of other exploratory studies of this 
general type.  The study is not powered to test non-inferiority or equivalency in immunogenicity 
among groups according to vaccination dosage and route of administration. 
  
 
 
 


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Rabies Vaccine  
RABVAX Version 6.0 20 MAY 2016 
  
Page 13 of 69 
 
Study Procedures: Group 1 (IM 3 doses) 
12 Subjects, 12 Visits, 13 Months Duration Per Subject 
 
Study Day 
-60 to 0 
0 
 
3 
 
7 
 
10 
 
14 
 
21 
24 
28 
77 
365 
 
372 
Visit Type 
Screening 
VAC 
BL 
BL 
VAC 
B L 
BL 
BL 
VAC  
BL 
 
BL 
 
BL 
BL 
BOOST 
BL 
BL 
Time windows (days) 
-60 to 0 
0 
 
+/-1 
 
+/-1 
 
+/-1 
 
+/-1 
 
+/-1 
+/-1 
+/-2 
+/-7 
+/-15 
 
+/-15 
Informed consent/IC review 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
Inclusion/exclusion criteria 
X 
X 
 
 
X 
 
 
 
X 
 
 
 
X 
 
Contraindications 
X 
X 
 
 
X 
 
 
 
X 
 
 
 
X 
 
Significant medical history 
X 
X 
 
 
X 
 
 
 
X 
 
 
 
X 
 
Physical exam 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
Targeted PE 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
X 
X 
 
X 
Vital signs 
X 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
X 
X 
 
X 
Urine pregnancy test 
*Females 
 
X* 
 
X* 
 
 
X* 
 
 
 
X* 
 
 
 
 
X* 
Concomitant therapy 
X 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
X 
X 
 
X 
Demography 
X 
 
 
 
 
 
 
 
 
 
 
 
Randomization 
 
X 
 
 
 
 
 
 
 
 
 
 
Blood Draw for  
Research Assays 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
X 
X 
 
X 
Vaccination 
 
X 
 
 
X 
 
 
 
X 
 
 
 
X 
 
AE and SAE 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
X 
 
X 
 
X 
 
X 
 
 
X 
 
 
 
 


Confidential/Proprietary Information 
Rabies Vaccine  
RABVAX Version 6.0 20 MAY 2016 
  
Page 14 of 69 
 
Blood Draw Table: Group 1 
Rabies vaccine Day 0, Day 7 and Day 21 
 
 
Day 
0 
3 
7 
10 
14 
 
21 
 
24 
 
28 
 
77 
 
365 
 
372 
 
 
VAC 
 
 
VAC 
 
 
 
 
VAC 
 
 
 
 
Boost 
 
Antibody 
Assays/ 
Proteomics  
4 x  
8.5 ml 
1 x 
 3 ml 
2 x 
 8.5 ml 
1 x 
3 ml 
2 x 
 8.5ml 
 
2 x  
8.5 ml 
 
1 x 
3 ml 
 
2 x 
 8.5 ml 
 
2 x  
8.5 ml 
 
4 x  
8.5 ml 
 
4 x  
8.5 ml 
CMI  
8 x 
 8.0 ml 
2 x 
 8.0 ml 
6x  
8.0 ml 
2 x  
8.0 ml 
5 x 
 8.0 ml 
 
5 x  
8.0 ml 
 
2 x 
 8.0 ml 
 
5 x 
 8.0 ml 
 
5 x  
8.0 ml 
 
8 x 
 8.0 ml 
 
3 x 
 8.0 ml 
Daily BV 
98 ml 
19 ml 
65  ml 
19 ml 
57 ml 
 
 
57 ml 
 
 
19 ml 
 
 
57 ml 
 
 
57 ml 
 
 
98 ml 
 
 
58 ml 
CBV (Total) 
98 ml 
117 ml 
182 ml 
201 ml 
258 ml 
 
 
 
 
315 ml 
 
 
 
 
 
334 ml 
 
 
 
 
391 ml 
 
 
 
 
448 ml 
 
 
 
 
546 ml 
 
 
 
 
604 ml 
Total  
(56 days) 
 
 
 
 
 
 
 
 
 
391 ml 
 
 
57 ml 
 
 
98 ml 
 
 
156 ml 
 


Confidential/Proprietary Information 
Rabies Vaccine  
RABVAX Version 6.0 20 MAY 2016 
  
Page 15 of 69 
 
 
 
 
Study Procedures: Group 2 (ID 3 doses) 
12 Subjects, 12 Visits, 13 Months Duration Per Subject 
 
Study Day 
-60 to 0 
0 
 
3 
 
7 
 
10 
 
14 
 
21 
24 
28 
77 
365 
 
372 
Visit Type 
Screening 
VAC 
BL 
BL 
VAC 
B L 
BL 
BL 
VAC  
BL 
 
BL 
 
BL 
BL 
BOOST 
BL 
BL 
Time windows (days) 
-60 to 0 
0 
 
+/-1 
 
+/-1 
 
+/-1 
 
+/-1 
 
+/-1 
+/-1 
+/-2 
+/-7 
+/-15 
 
+/-15 
Informed consent/IC review 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
Inclusion/exclusion criteria 
X 
X 
 
 
X 
 
 
 
X 
 
 
 
X 
 
Contraindications 
X 
X 
 
 
X 
 
 
 
X 
 
 
 
X 
 
Significant medical history 
X 
X 
 
 
X 
 
 
 
X 
 
 
 
X 
 
Physical Exam 
X 
 
 
 
 
 
 
 
 
 
 
 
Targeted PE 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
X 
X 
 
X 
Vital Signs 
X 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
X 
X 
 
X 
Urine pregnancy test 
*Females 
 
X* 
 
X* 
 
 
X* 
 
 
 
X* 
 
 
 
 
X* 
Concomitant therapy 
X 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
X 
X 
 
X 
Demography 
X 
 
 
 
 
 
 
 
 
 
 
 
Randomization 
 
X 
 
 
 
 
 
 
 
 
 
 
Blood Draw for  
Research Assays 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
X 
X 
 
X 
Vaccination 
 
X 
 
 
X 
 
 
 
X 
 
 
 
X 
 
AE and SAE 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
X 
 
X 
 
X 
 
X 
 
 
X 
 
 
 


Confidential/Proprietary Information 
Rabies Vaccine  
RABVAX Version 6.0 20 MAY 2016 
  
Page 16 of 69 
 
 
Blood Draw Table: Group 2 
Rabies vaccine Day 0, Day 7 and Day 21 
 
 
 
Day 
0 
3 
7 
10 
14 
 
21 
 
24 
 
28 
 
77 
 
365 
 
372 
 
 
VAC 
 
 
VAC 
 
 
 
 
VAC 
 
 
 
 
Boost 
 
Antibody 
Assays/ 
Proteomics  
4 x  
8.5 ml 
1 x 
3 ml 
2 x 
 8.5 ml 
1 x 
3 ml 
2 x  
8.5 ml 
 
2 x  
8.5 ml 
 
1 x  
3 ml 
 
2 x 
 8.5 ml 
 
2 x 
 8.5 ml 
 
4 x 
 8.5 ml 
 
4 x  
8.5 ml 
CMI  
8 x 
 8.0 ml 
2 x 
 8.0 ml 
6 x  
8.0 ml 
2 x  
8.0 ml 
5 x 
 8.0 ml 
 
5 x  
8.0 ml 
 
2 x 
 8.0 ml 
 
5 x 
 8.0 ml 
 
5 x  
8.0 ml 
 
8 x 
 8.0 ml 
 
3 x 
 8.0 ml 
Daily BV 
98 ml 
19 ml 
65 ml 
19 ml 
57 ml 
 
 
57 ml 
 
 
19 ml 
 
 
57 ml 
 
 
57 ml 
 
 
98 ml 
 
 
58 ml 
CBV (Total) 
98 ml 
117 ml 
182 ml 
201 ml 
258 ml 
 
 
 
 
315 ml 
 
 
 
 
334 ml 
 
 
 
 
391 ml 
 
 
 
 
448 ml 
 
 
 
 
546 ml 
 
 
 
 
604 ml 
Total 
(56 days) 
 
 
 
 
 
 
 
 
 
391 ml 
 
 
57 ml 
 
 
98 ml 
 
 
156 ml 
 
 
 


Confidential/Proprietary Information 
Rabies Vaccine  
RABVAX Version 6.0 20 MAY 2016 
  
Page 17 of 69 
 
Study Procedures: Group 3 (IM 2 doses) 
12 Subjects, 10 Visits, 13 Months Duration Per Subject 
 
Study Day 
-60 to 0 
0 
 
3 
 
7 
 
10 
 
14 
 
28 
 
77 
365 
372 
Visit Type 
Screening 
VAC 
BL 
 
BL 
VAC 
BL 
 
BL 
 
B L 
 
      BL 
 
BL 
BOOST 
BL 
 
BL 
Time windows (days) 
-60 to 0 
0 
 
+/- 1 
 
+/-1 
 
+/-1 
 
+/-1 
 
+/-2 
 
+/-7 
+/-15 
+/-15 
Informed consent/IC review 
X 
X 
 
 
 
 
 
 
 
 
 
 
Inclusion/exclusion criteria 
X 
X 
 
 
X 
 
 
 
 
 
 
X 
 
Contraindications 
X 
X 
     
      X 
 
 
 
 
 
 
X 
 
Significant medical history 
X 
X 
 
 
X 
 
 
 
 
 
 
X 
 
Physical Exam 
X 
 
 
 
 
 
 
 
 
 
Targeted PE 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
Vital Signs 
X 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
Urine pregnancy test 
*Females 
 
X* 
     X* 
 
 
      X* 
 
 
 
 
 
 
 
 
X* 
 
Concomitant therapy 
X 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
Demography 
X 
 
 
 
 
 
 
 
 
 
Randomization 
 
X 
 
 
 
 
 
 
 
 
Blood Draw for  
Research Assays 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
Vaccination 
 
X 
 
 
X 
 
 
 
 
 
 
X 
 
AE and SAE 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
X 
 
X 
 
 
 
 


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Rabies Vaccine  
RABVAX Version 6.0 20 MAY 2016 
  
Page 18 of 69 
 
Blood Draw Table: Group 3 
Rabies vaccine at Day 0 and Day 7 
 
 
 
Day 
0 
 
3 
7 
 
10 
14 
 
28 
 
77 
 
365 
 
372 
 
 
VAC 
 
VAC 
 
 
 
 
 
Boost 
 
Antibody 
Assays/ 
Proteomics  
4 x  
8.5 ml 
 
1 x  
3 ml 
2 x 
 8.5 ml 
 
1 x 
3 ml 
2 x  
8.5 ml 
 
2 x 
 8.5 ml 
 
2 x 
 8.5 ml 
 
4 x 
 8.5 ml 
 
4 x  
8.5 ml 
CMI  
8 x 
 8.0 ml 
 
2 x 
8.0 ml 
5 x  
8.0 ml 
 
2 x 
8.0 ml 
5 x 
 8.0 ml 
 
5 x 
 8.0 ml 
 
5 x  
8.0 ml 
 
8 x 
 8.0 ml 
 
3 x 
 8.0 ml 
Daily BV 
98 ml 
 
 
19 ml 
57 ml 
 
 
19 ml 
57 ml 
 
 
 
57 ml 
 
 
 
57 ml 
 
 
 
98 ml 
 
 
58 ml 
CBV (Total) 
98 ml 
 
 
 
 
117 ml 
174 ml 
 
 
 
 
193 ml 
250 ml 
 
 
 
 
307 ml 
 
 
 
364 ml 
 
 
 
462 ml 
 
 
 
520 ml 
Total (56 
days) 
 
 
 
 
 
 
307 ml 
 
57 ml 
 
98 ml 
 
156 ml 
 
 


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Study Procedures: Group 4 (ID 2 doses) 
12 Subjects, 10 Visits, 13 Months Duration Per Subject 
 
Study Day 
-60 to 0 
0 
 
3 
 
7 
 
10 
 
14 
 
28 
 
77 
365 
372 
Visit Type 
Screening 
VAC 
BL 
 
BL 
VAC 
BL 
 
BL 
 
B L 
 
      BL 
 
BL 
BOOST 
BL 
 
BL 
Time windows (days) 
-60 to 0 
0 
 
+/- 1 
 
+/-1 
 
+/- 1 
 
+/-1 
 
+/-2 
 
+/-7 
+/-15 
+/-15 
Informed consent/IC review 
X 
X 
 
 
 
 
 
 
 
 
 
 
Inclusion/exclusion criteria 
X 
X 
 
 
X 
 
 
 
 
 
 
X 
 
Contraindications 
X 
X 
     
      X 
 
 
 
 
 
 
X 
 
Significant medical history 
X 
X 
 
 
X 
 
 
 
 
 
 
X 
 
Physical Exam 
X 
 
 
 
 
 
 
 
 
 
Targeted PE 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
Vital signs 
X 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
Urine pregnancy test 
*Females 
 
X* 
     X* 
 
 
      X* 
 
 
 
 
 
 
 
 
X* 
 
Concomitant therapy 
X 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
Demography 
X 
 
 
 
 
 
 
 
 
 
Randomization 
 
X 
 
 
 
 
 
 
 
 
Blood Draw for  
Research Assays 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
 
X 
X 
X 
Vaccination 
 
X 
 
 
X 
 
 
 
 
 
 
X 
 
AE and SAE 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
 
X 
 
X 
 
 
 
 
 


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Blood Draw Table: Group 4 
Rabies vaccine at Day 0 and Day 7 
 
 
Day 
0 
 
3 
7 
 
10 
14 
 
28 
 
77 
 
365 
 
372 
 
 
VAC 
 
VAC 
 
 
 
 
 
Boost 
 
Antibody 
Assays/ 
Proteomics  
4 x  
8.5 ml 
 
1 x 
3 ml 
2 x 
 8.5 ml 
 
1 x 
3 ml 
2 x  
8.5 ml 
 
2 x 
 8.5 ml 
 
2 x 
 8.5 ml 
 
4 x 
 8.5 ml 
 
4 x  
8.5 ml 
CMI  
8 x 
 8.0 ml 
 
 
2 x 
8.0 ml 
5 x  
8.0 ml 
 
 
2 x 
8.0 ml 
5 x 
 8.0 ml 
 
5 x 
 8.0 ml 
 
5 x  
8.0 ml 
 
8 x 
 8.0 ml 
 
3 x 
 8.0 ml 
Daily BV 
98 ml 
 
 
19 ml 
57 ml 
 
 
19 ml 
57 ml 
 
 
57  ml 
 
 
57 ml 
 
 
98 ml 
 
 
58 ml 
CBV (Total) 
98 ml 
 
 
 
 
117 ml 
174 ml 
 
 
 
 
193 ml 
250 ml 
 
 
 
 
307 ml 
 
 
 
 
364  ml 
 
 
 
 
462 ml 
 
 
 
 
520 ml 
Total (56 
days) 
 
 
 
 
 
 
307 ml 
 
57 ml 
 
98 ml 
 
156 ml 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


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Study Procedures: Group 5 (control) 
6 Subjects, 4 Visits, 7 Day Duration Per Subject 
 
 
 
 
 
 
          Study Day 
-60 to 0 
0 
 
3 
 
7 
Visit Type 
Screening 
VAC 
BL 
 
BL 
 
B L 
Time windows (days) 
-60 to 0 
0 
 
+/-1 
 
+/-1 
Informed consent/IC review 
X 
X 
 
 
 
 
Inclusion/exclusion criteria 
X 
X 
 
 
 
 
Contraindications 
X 
X 
     
 
 
Significant medical history 
X 
 
 
 
 
 
Physical Exam 
X 
 
 
 
Targeted PE 
 
X 
 
X 
 
X 
Vital signs 
X 
X 
 
X 
 
X 
Urine pregnancy test 
*Females 
 
X* 
     X* 
 
 
 
Concomitant therapy 
X 
X 
 
X 
 
X 
Demography 
X 
 
 
 
Randomization 
 
X 
 
 
Blood Draw for  
Research Assays 
 
X 
 
X 
 
X 
Vaccination 
 
X 
 
 
 
 
AE and SAE 
 
 
X 
 
 
X 
 
 
X 


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Blood Draw Table: Group 5 
Human Serum Albumin at Day 0 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Day 
0 
3 
7 
 
 
VAC 
 
 
 
Proteomics  
1 x  
3 ml 
1 x 
3 ml 
1 x  
3 ml 
Daily BV 
3 ml 
3 ml 
3 ml 
CBV (Total) 
3 ml 
6 ml 
9 ml 
 
 
 
 


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Study Procedures: Group 6 (control) 
6 Subjects, 4 Visits, 7 Day Duration Per Subject 
 
Study Day 
-60 to 0 
0 
 
3 
 
7 
Visit Type 
Screening 
VAC 
BL 
 
BL 
 
B L 
Time windows (days) 
-60 to 0 
0 
 
+/-1 
 
+/-1 
Informed consent/IC review 
X 
X 
 
 
 
 
Inclusion/exclusion criteria 
X 
X 
 
 
 
 
Contraindications 
X 
X 
     
 
 
Significant medical history 
X 
X 
 
 
 
 
Physical Exam 
X 
 
 
 
Targeted PE 
 
X 
 
X 
 
X 
Vital signs 
X 
X 
 
X 
 
X 
Urine pregnancy test 
*Females 
 
X* 
     X* 
 
 
 
Concomitant therapy 
X 
X 
 
X 
 
X 
Demography 
X 
 
 
 
Randomization 
 
X 
 
 
Blood Draw for  
Research Assays 
 
X 
 
X 
 
X 
Vaccination 
 
X 
 
 
 
 
AE and SAE 
 
 
X 
 
 
X 
 
 
X 
 
 
 
 
 


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Blood Draw Table: Group 6 
Human Serum Albumin at Day 0 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Day 
0 
3 
7 
 
 
VAC 
 
 
 
Proteomics  
1 x  
3 ml 
1 x 
 3 ml 
1 x  
3 ml 
Daily BV 
     3 ml 
3 ml 
3 ml 
CBV (Total) 
3 ml 
6ml 
9 ml 


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List of Terms and 
Abbreviations 
Ab 
antibody 
ADR 
adverse drug reaction 
AE 
adverse event 
AESI 
adverse event of special interest 
AR 
adverse reaction 
BL 
blood sample visit 
C 
Celsius 
CDM 
Clinical Data Management 
CI 
confidence interval 
CIOMS 
Council for International Organizations of Medical Sciences 
CMI 
Cell Mediated Immunity 
CRA 
Clinical Research Associate 
CSTE 
Council of State and Territorial Epidemiologists 
D 
day 
eCRF 
electronic case report form  
EDC 
electronic data capture 
ELISA 
enzyme linked immuno sorbent assay 
F 
Fahrenheit 
FAS 
full analysis set 
GCI 
Global Clinical Immunology 
GCP 
Good Clinical Practices 
GM 
geometric mean  
GMT 
geometric mean titer 
GMTR 
geometric mean titer ratio 
HIV 
human immunodeficiency virus 


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   ICF 
informed consent form 
   ID                                      intradermal 
IM           
intramuscular 
IRB 
Institutional Review Board 
   M 
month 
   NR 
not reportable 
OD 
optical density 
PBMC 
peripheral blood mononuclear cell 
PDF 
portable document format 
PPAS 
per-protocol analysis set 
Product 
Rabies vaccine 
QA 
quality assurance 
R&D 
Research and Development 
RCDC 
reverse cumulative distribution curve 
RFFIT 
rapid fluorescent focus inhibition test 
SADR 
Serious Adverse Drug Reaction 
SAE 
serious adverse event 
SC 
subcutaneous(ly) 
SOP 
standard operating procedure 
UAR 
unexpected adverse reaction 
US 
United States 
V 
visit 
VAC 
vaccination (with Rabies vaccine) 
WRAIR 
Walter Reed Army Institute of Research 


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1 
Introduction 
 
This is an exploratory vaccine trial to evaluate immunogenicity of a non-licensed dosing schedule 
and route of administration for a currently FDA licensed rabies vaccine for pre-exposure 
prophylaxis against rabies infection.  The goal of this study is to characterize the immune response 
and persistence of immunity to a shortened dose schedule and intradermal (ID) administration, 
relative to the current licensed dosing schedule of the rabies vaccine (3 dose (0, 7, 21 days) IM). 
Rabies virus is endemic throughout the world due to high rates of both wild and domestic animal 
rabies and the risk to deployed military in endemic areas is considerable. Currently the commonly 
supported pre-exposure prophylaxis regimen for rabies, in the United States is comprised of three, 
1.0 ml intramuscular (IM) injections of the human diploid cell vaccine (HDCV) or purified chick 
embryo cell (PCEC) rabies vaccine on days 0, 7, and 21 or 28.  Modified, two and three dose 
schedules of intradermal (ID) injections of 0.1 ml of HDCV and PCEC are the standard outside the 
US. These two and three dose intradermal schedules share a similar safety and immunogenicity 
profile to intramuscular vaccinations and are easily boosted at one year after vaccination. A death, 
this year, from rabies, of a US Soldier returned from Afghanistan underscores the importance of 
rabies pre-exposure prophylaxis for soldiers and the need to evaluate the safest, most effective 
means of vaccinating large deploying forces.  While the current three dose, 1 ml IM rabies series is 
effective, a shortened, equally effective vaccination series with significantly smaller dose per 
injection would greatly improve the logistics and cost associated with universal or even targeted 
coverage of deploying soldiers.  Evaluation of a shorter, smaller-dose, pre-exposure vaccination 
series for rabies is the goal of this study. 
 
1.1 
Background of Rabies 
 
Rabies virus is a RNA virus, serotype 1 of 7 serotypes of the genus Lyssavirus, family Rhabdovirus 
(1). The lyssaviruses including rabies virus are found in a number of mammalian species and 
transmitted to humans as a zoonotic disease.  The primary animal vector varies geographically.  In   
North America: skunks, raccoons, foxes, bats are the primary animal vector; Western Europe: foxes 
and bats; Eastern Europe: foxes and dogs; Latin America: dogs and bats; Caribbean: mongooses; 
Africa: dogs, jackals, mongooses, and foxes; Asia: dogs, cats, monkeys, mongooses, and arctic 
foxes.  The outer part of the virus protein contains the matrix protein and a glycoprotein on which 
are located the epitopes that induce neutralizing antibodies, components of the current vaccine, and 
responsible for protection from infection.  Rabies is a global health problem with 55,000 deaths 
occurring every year (2).  It is highly endemic in most areas of the world (figure 1).  Considering 
that there is a highly effective pre and post-exposure prophylaxis regimen to rabies exposure, the 
global health problem is one of an access to care and effective rabies vaccines. 
 
 
 
 
 
 
 


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Figure 1. Global risk of rabies (World Health Organization, 
http://www.who.int/rabies/rabies_maps/en/index.html ) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
1.2 
Background of the Licensed Product       
 
A vaccine against rabies was the first vaccine to have been developed by Louis Pasteur; it was 
grown in and purified fromrabbit spinal cords.  The rabies vaccine that is currently available in the 
U.S. is derived from cell cultures, either a human diploid cell vaccine (HDCV) or as a purified 
chick embryo cell vaccine (PCECV) (1).  Imovax Rabies (HDCV for pre or post-exposure) was 
developed by Sanofi Pasteur and licensed in1980.  RabAvert (PCECV for pre or post-exposure) was 
developed by Novartis and licensed in 1997. Table 1 demonstrates the current regimens for both pre 
and post-exposure prophylaxis with the rabies vaccine. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


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Table 1. Regimens for pre-exposure and post-exposure vaccination with rabies vaccines (1). 
 
The current rabies vaccines produce antibody response in 100% of individuals with durable 
immunity for years.  In a prospective study over 10 years of follow-up, an antibody titer of 130 IU 
following vaccination indicated prolonged seropositivity with titers of 0.5 IU or greater considered 
protective (3).  Pre-exposure vaccination is recommended for people who will be exposed to rabies 
virus either in the laboratory, environment or by occupation.  It is recommended for travelers or 
global health workers who may have potential exposure (1).  Reactions to the current rabies 
vaccines are mild with local pain, erythema, and swelling at the injection site.  Systemic reactions 
are less frequent with headache and malaise (1).  Guillain-Barré syndrome has been reported as a 
rare event following vaccination but its association with vaccination is uncertain (1).  Allergic 
reactions were thought to result from the formation of antibodies to the human albumin stabilizer 
that is chemically altered by beta-propiolactone. Systemic reactions to newer vaccines with 
significantly lower concentrations of human albumin are rare (11).  The current licensed delivery of 
the rabies vaccine is by the IM route. Due to the high cost of cell cultured vaccines strategies have 
been investigated to reduce the number of vaccine doses at an equivalent immune response by ID 
administration (4). Intradermal vaccination is thought to produce a better immune response as 
compared to IM delivery due to the antigen processing of Langerhans’ cells of the epidermis.  The 
ID dose is one-fifth of a dose of an IM delivery.  
 
 
1.3 
Potential Benefits to Subjects 
 
Subjects Injected with Rabies Vaccine 
 
Subjects will benefit from the rabies vaccine by developing protective antibody to the rabies virus.   
 
  
1.4 
Potential Risks to Subjects 
 
Subjects Injected with Rabies Vaccine 
 
Reactions to the current rabies vaccines as previously noted are mild with local pain, erythema, and 
swelling at the injection site.  Systemic reactions are less frequent and include headache and malaise 
(1).   
 


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All subjects 
 
Potential risks may also include the unwanted effects of blood sampling, which may include 
tenderness or bruising at the spot where blood was drawn. Fainting or dizziness can occur after a 
blood draw, but this is uncommon. 
 
 
1.5 
Rationale 
 
Antibody Assays:  The rabies virus envelope glycoprotein G is the correlate for protective 
immunity and by World Health Organization (WHO) guidelines, an antibody titer for protection 
should be greater than or equal to 0.5 IU/ml. Several studies have demonstrated the high sensitivity 
and specificity of the Bio-Rad, enzyme-linked immunosorbent assay (ELISA), PLATELIA 
RABIES II® and correlation to the current reference method, rapid fluorescent focus inhibition test 
(RFFIT) (5, 6). The Bio-Rad Platelia Rabies II assay will be used to assess subjects’ immune 
response to rabies vaccine.  
Proteomics:  Proteomics is the analysis of the expression, localizations, functions, and interactions 
of a set of proteins expressed by the genetic material of an organism under a given set of 
environmental conditions.  This technique offers a novel method to evaluate clinical samples from 
experimental vaccination for identification of novel biomarkers and correlates of immunogenicity. 
For this protocol, the specific aim is to identify subtle changes in the immune response associated 
with rabies vaccination given at various dosing intervals and both ID and IM. It is likely this 
analysis can better characterize the immune response in individuals whose titer drops below 0.5 
IU/mL and help determine if they are still protected as indicated by the rapid, robust return of high 
antibody titers with a single booster dose.  The iTRAQ (isobaric technique for relative and absolute 
quantitation) proteomic method allows the quantitative profiling of proteins from complex mixtures 
and is referred to as multi-dimensional protein identification technology (MudPIT) (13-15) (7-9). 
Upstate Medical University Proteomics Core facility will be used to test samples and consists of an 
LTQ Orbitrap XL mass spectrometer (16-18). The core proteomics facility employs both iTRAQ 
labeling and 2D DIGE methods of sample preparation for protein isolation and identification. Ettan 
Spot Picker, Decyder 2D differential analysis software and Typhoon 9410 variable mode imager 
are available for the 2D DIGE samples. For this proposal, iTRAQ labeling will be conducted 
according to the manufacturer’s protocol (Applied Biosystems). Initially, samples will undergo 
immunodepletion, using eluting columns to remove high abundance proteins that may overwhelm 
unique peptide signatures.  Briefly, 100 µg of protein in 20 µl is denatured and reduced with 
denaturant (1µl) and reducing agent (2 µl) supplied with the kit. The samples are incubated for 1 
hour at 60o C and cysteine blocking agent is added (1µl) followed by incubation at room temp for 
10 min. The samples are digested with trypsin at 37o C overnight and then processed by adding 
iTRAQ reagents. The samples are mixed and incubated another hour at room temp. The separation 
and mass spectral analysis will be conducted as described (19). The labeled 4 samples are pooled 
together, diluted 1:10 with a cation exchange buffer A and are fractionated by SCX HPLC. The 
sample is loaded in buffer A and eluted with a gradient of 0-350 mM KCl over 40 min at 0.2 
ml/min. Twenty fractions are collected and then desalted with a C18 zip tip prior to LC-MS/MS. 
Samples will be processed with a Orbitrap XL FT LC-MS-MS system. The fractions are 


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individually injected by autosampler onto a 5 µm C18 reverse phase column. The samples are 
eluted using a buffered gradient for 5 min. Nanospray data for MS and MS/MS are analyzed with 
ProQuant software as described (19) (10).  
Cell Mediated Immunity (CMI) 
We propose to monitor both the quality and quantity of immune activation immediately after 
vaccination as well the quality and quantity of the subsequently generated adaptive immunity in 
response to booster immunization with the RABVAX vaccine. Blood will be obtained for evaluation 
of cell-mediated immune responses to vaccination. At each time point blood will be collected in CPT 
tubes and PBMCs will be separated. The PBMCs will be stored frozen at -130C° or below until 
analyzed at SUNY Upstate Medical University or shipped to WRAIR for analysis. CMI assays will 
include ELISPOT assays and flow cytometry intracellular cytokine staining (ICS). Cryopreserved 
PBMC will be tested in batched format to determine the presence and frequency of vaccine 
responders and the magnitude of the immune response using IFN-g spot-forming units as a read-out. 
Assays for T cell function will test at a minimum for IFN-γ production, however additional analysis 
for production of other cytokines such as TNF-α and IL-2, as well as the generation of short and long 
term effector and memory T cell populations can be performed in an identical manner using 
multiparameter flow cytometry. Flow cytometry will be used to comprehensively assess cellular 
activation and maturation in all lymphoid and myeloid cell lineages in PBMC, determine the 
functional profile of antigen-specific cells, and directly phenotype the antigen-specific cells. 
 
2     Objectives 
 
2.1     Primary Objectives 
 
Immunogenicity 
1) To describe percentage of subjects achieving a protective  humoral immune response to 
each of the vaccine dosing schedules at day 28 and day 365 after first vaccine dose, and 7 
days after a booster given at one year (372 days) 
    
                        The endpoints for the primary objective(s) are presented in Section 16.1.  
 
 
    2.2    Secondary Objectives 
 
   
Immunogenicity 
 
1) To describe the persistence of the humoral immune response to each of the rabies 
vaccine routes 12 months post dose 1 
 
 
 
 
 
 


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2.3    Exploratory Objectives 
 
Immunogenicity 
1) Describe the cell-mediated immune response to rabies vaccination in groups 1-4. 
 
2) Proteomic characterization of the host response before and after each vaccination to 
identify biomarkers and correlates of immunogenicity. 
 
3 
Investigators and Trial Organization 
 
This trial will be conducted at Upstate Medical University (Upstate) in Syracuse, NY, US. The 
Principal Investigator and sub-investigators are from the Center for Global Health and 
Translational Science at Upstate Medical University. The trial team will be coordinated by Dr. 
Mark E. Polhemus.  
 
 
Laboratory 
 
 Serology or antibody titer by RFFIT assessments will be performed by the Rabies Division, Kansas State 
University Laboratory. Exploratory assays will be performed at Division of Virology, Walter Reed 
Army Institute of Research, and the Department of Microbiology and Immunology, Upstate 
Medical University.  Additional laboratories or contractors may be utilized in the development or 
conduct of some research assays.  
  
4 
Independent Ethics Committee / Institutional Review Board 
 
Before the inclusion of the first subject, this protocol, the informed consent form(s) (ICF), 
subject recruitment procedures, and any other written information to be provided to subjects 
must be approved by Upstate’s Institutional Review Board (IRB). 
 
In accordance with Good Clinical Practice (GCP) and local regulations, each Investigator and / or 
the Sponsor are / is responsible for obtaining this approval before the start of the trial.  If the 
protocol is subsequently amended, approval must be re-obtained for each amendment. Copies of 
these approvals, along with information on the type, version number, and date of document, and 
the date of approval, must be forwarded by the Investigator to the Sponsor.  
 
The Investigator will submit written summaries of the status of the trial to the IRB annually, or 
more frequently if requested. All serious adverse events (SAEs) occurring during the trial that are 
related to vaccination will be reported by the Investigator to the IRB, according to IRB policy. 
 
 
  5 
Trial Design 
 
This will be a randomized, open-label, single center study conducted in 60 healthy adults aged ≥ 18 
years to ≤ 60 years in the US. 
 
Rabies vaccine will be administered to subjects in the four vaccination groups as follows: 
 
 
 


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Table 2: Schedules of vaccine administration and number of subjects by group 
 
Vaccine 
Group 
 
Day 
Number per Group 
0 
7 
 
21 
365 
1 
IM 
IM 
 
IM 
IM 
12 
2 
ID 
ID 
 
ID 
IM 
12 
3 
IM 
IM 
 
IM 
12 
4 
ID 
ID 
 
IM 
12 
5 
IM 
control 
 
 
 
6 
5 
ID 
control 
 
 
 
6 
 
 
Subjects will be randomly assigned to one of the six groups through a computer generated process. 
A standard permuted-block design will be used for randomization/allocation in order to avoid 
undesirable difference in the numbers of subjects assigned to each group. 
 
Blood samples will be taken at each visit for research assays.  Assays include evaluation of 
antibody assays, proteomics, evaluation and Cell Mediated Immunity.  See section 5.3.   
 
In all groups additional blood samples may be taken if required to assess AEs or abnormal 
laboratory findings. Additional biological samples (e.g., body fluid, tissue samples) may be 
collected in the event of an AE, or SAE. See section 14.1.2. 
 
Clinical site personnel will record immediate unsolicited AEs that occur within 30 minutes after 
receiving the Rabies vaccine. The occurrence of SAEs will be collected throughout the study. 
  
5.1 
Justification of the Trial Design 
 
This exploratory study is designed to be a broad evaluation of the immune response, in healthy adults, 
to vaccination with rabies vaccine across a variety of vaccination scenarios.  The goal is to 
characterize the host response in four vaccination schedules. 
Evaluation will be focused on antibody response to trial regimens in relation to using the 
accepted rabies vaccination schedule. The goal is to evaluate host response to the delivery of a 
licensed vaccine using an ID delivery and a 2 dose schedule compared to the recommended 
vaccination schedule of 3 doses by IM delivery, using descriptive statistical methods and 
exploratory data analysis. 
The placebo (human serum albumin) groups will have proteomic analyses performed. This will 
show any differences in the proteomic responses that are due to the mechanical differences in the 
different routes of administration. 
 
 
 
 


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5.2 
Trial Plan 
 
Eligible subjects will be identified and recruited. Each subject must sign and date the informed 
consent form before any procedure or treatment associated with the study is performed. 
 
Depending on group assignment, subjects will receive vaccination in accordance with the schedule 
outlined in Table 2 above. There will be follow-up visits for review of adverse events and collection 
of blood samples.  
 
Blood samples will be collected at Day 0 and at various time points throughout the study to test 
the immune response to rabies. Additional blood samples will be collected at different time points 
to evaluate the immune response to rabies vaccination. Additional biological samples (e.g. body 
fluids, tissue samples) may be collected in the event of an SAE.         .  
 
5.3 
Visit Procedures 
 
Each of the six vaccination groups will each have a schedule of study procedures. These study 
procedures are described for each group separately below. Although the schedule of visits is 
different for each study group, the basic types of visit are the same and the same procedures will 
be followed. 
 
VAC: vaccination visit at which the rabies vaccine is administered. 
 
BL: blood sample visit at which a blood sample is obtained. These BL visits will occur at 
designated days or months after the previous administration of vaccine. If vaccination occurs on 
the same day, the blood sample will be obtained before vaccination. 
All screening assessments must be completed within 60days prior to vaccination.  
Screening numbers will be assigned when the subject signs the consent document starting with 
screening #100, 101, 102, etc.  When the subject has completed the screening visit and is eligible 
and willing to continue participation, they will be randomized according to the randomization 
process outlined in section 11.   Screen-fails will not be recorded in the data management system 
for the study. During the screening period, the following procedures and assessments will be carried 
out for each subject to determine their eligibility for participation in the study. 
• 
Obtain informed consent 
• 
Obtain medical history 
• 
Record concomitant medications 
• 
Record demographics  
• 
Complete physical examination 
• 
Review of inclusion and exclusion criteria  
• 
Vital signs  
• 
Urine pregnancy test (females of child bearing potential)  
 
On Day 0, when the subject arrives for his/her first vaccination, the following events will occur 
before randomization: 
1) Review informed consent 
2) Review inclusion and exclusion criteria to confirm eligibility. 
3) Review pertinent medical history Section 6.6 of the subject and any changes in medical 
history since screening.   
4) Conduct a directed physical examination, as required Section 6.6, measure the subject’s 


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temperature, and obtain vital signs.  
5) For all women, obtain a urine sample and perform a urine pregnancy test prior to 
vaccination.  Do not vaccinate if the test result is positive. 
6) Review the subject’s concomitant medications.   
7) Record information in the source document.  Concomitant medications and therapy will 
be recorded in the source document. 
 
If the subject is eligible for vaccination they will be randomized to one of the six groups 
and then vaccination will begin on one of the following schedules, depending on 
randomization.   
 
Group 1 
 
Vacc 
Group 
 
Day 
No. per Group 
00 
07 
     
    21 
365 
1 
IM 
IM 
IM 
IM 
12 
 
 
Day 0 Rabies Vaccination #1 
 
1) Confirm subject is randomized to Group 1. 
2) Review inclusion/exclusion criteria, including pregnancy test for females.  
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intra-muscular injection in the deltoid).  
9) Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
 
Day 3 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3)  Targeted physical exam. 
4) Concomitant medication review. 
5) Unsolicited AE/SAE assessment. 
 
Day 7 Rabies Vaccination #2 
 
1) Confirm subject is randomized to Group 1. 
2) Review inclusion/exclusion criteria, including pregnancy test for females 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 


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6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intra-muscular injection in the deltoid).  
9) Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
10) AE/SAE assessment. 
 
Day 10 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5)  AE/SAE assessment. 
 
Day 14 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 21 Rabies Vaccination #3 
 
1) 
Confirm subject is randomized to Group 1. 
2) 
Review inclusion/exclusion criteria, including pregnancy test for females 
3) 
Targeted physical exam. 
4) 
Concomitant medication review 
5) 
Review medical history. 
6) 
Obtain vital signs  
7) 
Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) 
Inject rabies vaccine (intra-muscular injection in the deltoid).  
9) 
Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
10) 
AE/SAE assessment. 
 
Day 24 post vaccination  
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 


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Day 28 post vaccination  
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) Unsolicited AE/SAE assessment. 
 
Day 77 post vaccination  
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 365 Rabies Boost  
 
1) Confirm subject is randomized to Group 1. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intra-muscular injection in the deltoid).  
9) Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
10) AE/SAE assessment. 
 
Day 372 post vaccination  
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
 
 
 
 
 
 
 
 
 


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Group 2 
 
 
Vacc 
Group 
 
Day 
No. per Group 
00 
07 
 
21 
365 
2 
ID 
ID 
 
ID 
IM 
12 
 
 
Day 0 Rabies Vaccination #1 
 
1) Confirm subject is randomized to Group 2. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intradermal injection in the forearm).  
9) Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
 
Day 3 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
Day 7 Rabies Vaccination #2 
 
1) Confirm subject is randomized to Group 2. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intradermal injection in the forearm). Observe subject for 30 
minutes, and record any adverse reaction (s) in the source document and the eCRF. 
9) AE/SAE assessment. 
 
 
 
 


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Day 10  post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Directed history and physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
Day 14  post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
Day 21 Rabies Vaccination #3 
 
1)   Confirm subject is randomized to Group 2. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intradermal injection in the forearm). Observe subject for 30 
minutes, and record any adverse reaction (s) in the source document and the eCRF. 
9) AE/SAE assessment. 
 
Day 24 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 28 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
 
 


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Day 77 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 365 Rabies Boost 
 
1)   Confirm subject is randomized to Group 2. 
2) 
Review inclusion/exclusion criteria, including pregnancy test for females. 
3) 
Targeted physical exam. 
4) 
Concomitant medication review 
5) 
Review medical history. 
6) 
Obtain vital signs  
7) 
Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) 
Inject rabies vaccine (intra-muscular injection in the deltoid). Observe subject for 30 
minutes, and record any adverse reaction (s) in the source document and the eCRF. 
9) 
AE/SAE assessment. 
 
Day 372 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


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Group 3  
 
 
 
 
 
 
 
 
 
Day 0 Rabies Vaccination #1 
 
1) Confirm subject is randomized to Group 3. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intra-muscular injection in the deltoid).  
9) Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
 
Day 3 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted  physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 7 Rabies Vaccination #2 
 
1) Confirm subject is randomized to Group 2. 
2) 
Review inclusion/exclusion criteria, including pregnancy test for females. 
3) 
Targeted physical exam. 
4) 
Concomitant medication review 
5) 
Review medical history. 
6) 
Obtain vital signs  
7) 
Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) 
Inject rabies vaccine (intra-muscular injection in the deltoid).  
Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
9)   AE/SAE assessment 
 
 
 
Vacc 
Group 
 
        Day             No. per Group 
00 
 
07 
365 
 
3 
IM 
 
IM 
IM 
12 


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Day 10 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted  physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 14 post vaccination 
 
1)   Obtain vital signs. 
2)   Obtain blood samples for antibody assays, proteomics and CMI. 
3)   Targeted  physical exam. 
4)   Concomitant medication review. 
5)   AE/SAE assessment. 
 
Day 28 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 77 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted  physical exam. 
4) Concomitant medication review. 
5) AE/SAE  assessment. 
 
Day 365 Rabies Boost 
 
1) Confirm subject is randomized to Group 3. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intra-muscular injection in the deltoid). Observe subject for 30 
minutes, and record any adverse reaction (s) in the source document and the eCRF. 
9) AE/SAE assessment. 
 
 
 
 


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Day 372 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
Group 4 
 
 
 
 
 
 
 
 
 
Day 0 Rabies Vaccination #1 
 
1) 
Confirm subject is randomized to Group 4. 
2)    Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intradermal injection in the forearm).  
9)   Observe subject for 30 minutes, and record any adverse reaction(s) in the source          
document and the eCRF. 
 
Day 3 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted  physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 7 Rabies Vaccination #2 
 
1) Confirm subject is randomized to Group 4. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
Vacc 
Group 
 
Day             No. per Group 
00 
 
07 
365 
 
4 
ID 
 
ID 
IM 
12 


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be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intradermal injection in the forearm). Observe subject for 30 
minutes, and record any adverse reaction (s) in the source document and the eCRF. 
9) AE/SAE assessment. 
 
Day 10 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted  physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 14 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 28 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 77 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
Day 365 Rabies Boost 
 
1) Confirm subject is randomized to Group 4. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for antibody assays, proteomics, CMI.  The blood samples must 
be obtained before vaccination from the arm opposite to the one to be used for 
vaccination. 
8) Inject rabies vaccine (intra-muscular injection in the deltoid). Observe subject for 30 


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minutes, and record any adverse 
reaction (s) in the source document 
and the eCRF. 
9) 
Observe subject for 30 minutes, 
and record any adverse reaction(s) in 
the source document and the eCRF. 
10) AE/SAE assessment. 
 
Day 372  post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for antibody assays, proteomics and CMI. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
Group 5 
 
 
 
 
 
 
 
 
Day 0 Control Vaccination  
 
1)    Confirm subject is randomized to Group5. 
2)    Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for, proteomics. The blood samples must be obtained before 
vaccination from the arm opposite to the one to be used for vaccination. 
8) Inject human serum albumin (intramuscular injection in the deltoid).  
9) Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
10) AE/SAE assessment. 
 
Day 3 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for proteomics. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
Vacc 
Group 
 
Day             No. per Group 
00 
 
03 
07 
 
5 
IM 
 
 
 
6 


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Day 7 post vaccination 
 
1) Obtain vital signs. 
2) Obtain blood samples for proteomics. 
3) Targeted physical exam. 
4) Concomitant medication review. 
5) AE/SAE assessment. 
 
 
Group 6 
 
Vacc 
Group 
 
Day             No. per Group 
00 
 
 03 
07 
 
6 
ID 
 
 
 
6 
 
Day 0 Control Vaccination  
 
1) Confirm subject is randomized to Group 6. 
2) Review inclusion/exclusion criteria, including pregnancy test for females. 
3) Targeted physical exam. 
4) Concomitant medication review 
5) Review medical history. 
6) Obtain vital signs  
7) Obtain blood samples for proteomics.  The blood samples must be obtained before 
vaccination from the arm opposite to the one to be used for vaccination. 
8) Inject human serum albumin (intradermal injection in the forearm).  
9) Observe subject for 30 minutes, and record any adverse reaction(s) in the source 
document and the eCRF. 
 
Day 3 post vaccination 
 
1) Obtain vital signs. 
2)   Obtain blood samples for proteomics. 
3)   Targeted physical exam. 
   4) Concomitant medication review. 
5)   AE/SAE assessment. 
 
Day 7 post vaccination 
 
1)  Obtain vital signs.   
2) Obtain blood samples for proteomics. 
   3)   Targeted physical exam. 
   4)   Concomitant medication review. 
   5)   AE/SAE assessment. 
 


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5.4 Planned Trial Calendar 
 
The following dates are approximate. The actual dates may differ as, for example, the trial will 
not start until all the appropriate regulatory and ethical approvals have been obtained. 
 
February 2014: IRB Submission 
Planned trial period - FVFS (first visit, first subject) to LCLS (last contact, last subject): October 
2014, February 2016 
October 2014: First subject, first vaccination 
January 2015: Last subject, last vaccination 
February 2016: Last contact, last subject/Final study visit 
July 2016: Assays Complete 
January 2017: Final clinical study report submission 
 
 
6 
Enrollment and Retention of Trial Population 
 
 
6.1 
Recruitment Procedures 
 
Subjects will be recruited for participation in this clinical trial. The site will ensure that any 
advertisements used to recruit subjects (letters, pamphlets, posters, etc.) are submitted to the IRB 
for approval. 
 
6.2 
Informed Consent Procedures 
 
Informed consent is the process by which a subject voluntarily confirms his or her willingness to 
participate in a particular trial. Informed consent must be obtained before any study procedures 
are performed. The process is documented by means of a written, signed, and dated ICF.  In 
accordance with GCP, prior to signing and dating the consent form, the subject must be informed 
by appropriate study personnel about all aspects of the trial that are relevant to make the decision 
to participate, and must have sufficient time and opportunity to ask any questions. 
 
 
Any change to the content of the ICF must be approved by the IRB prior to the form being used. 
 
If new information becomes available that may be relevant to the subject’s willingness to continue 
participation in the trial, this will be communicated to him/her in a timely manner. Such 
information will be provided via a revised ICF or an addendum to the original ICF. 
 
Informed consent forms will be photocopied. The original will be kept by the Investigator, and 
the copy will be kept by the subject. 
 
Documentation of the consent process should be recorded in the source documents. 
 
6.3 
Screening Criteria 
 
There are no screening criteria other than the inclusion and exclusion criteria. 
 


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  6.4 
Inclusion Criteria 
 
Subjects must fulfill all of the following criteria in order to be eligible for trial enrollment: 
1) Aged ≥ 18 to ≤ 60 years on the day of inclusion 
 
2) Informed consent form has been signed and dated 
 
3) Able to attend all scheduled visits and to comply with all trial procedures 
 
4) Subject in good health, based on medical history and physical examination 
 
a “18 to 60 years” means from the day of the 18th birthday to the day before the 60th  birthday 
 
  6.5        Exclusion Criteria 
 
Subjects fulfilling any of the following criteria are to be excluded from trial enrollment: 
 
1. Subject is pregnant, or lactating, or of childbearing potential (to be considered of non-
childbearing potential, a female must be post- menopausal for at least 1 year, surgically sterile, or 
using an effective method of contraception or abstinence from at least 4 weeks prior to the first 
vaccination and until at least 4 weeks after the last vaccination 
  
2.   Participation in the 4 weeks preceding the first trial vaccination, or planned participation during 
the present trial period, in another clinical trial investigating a vaccine, drug, medical device, or 
medical procedure.  
 
3. Previous history of receiving the rabies vaccine 
 
4.   Previous history of receiving rabies immune globulin 
 
5.   Any major psychiatric disorder, such as severe depression, severe anxiety disorder, psychosis, 
schizophrenia, other major psychiatric disorders, or seizures. History of mild depression or 
anxiety disorder that is well controlled is not an exclusion criteria. 
 
6.  Any history of cardiac arrhythmias, such as:  bradycardia, tachycardia, heart block, SVT, PAC, 
VT, VF, or any other conduction abnormalities. 
 
7.   Use of any immunosuppressive drug, including topical steroids of potency groups I, II or III 
within 30 days of the study period. 
 
8.  Any immunosuppressive disorder, such as HIV infection, common variable immunodeficiency, 
active cancers or chemotherapy. 
 
9.   History of renal insufficiency or requiring dialysis.  
 
10.   Have any condition that would, in the opinion of the site investigator, place the subject at an 
unacceptable risk of injury or render the subject unable to meet the requirements of the protocol. 
 


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11.  Identified as an employee of the Investigator or study center, with direct involvement in the 
proposed study or other studies under the direction of that Investigator or study center, as well as 
family members (i.e., immediate, husband, wife and their children, adopted or natural) of the 
employee or the Investigator. 
 
12. Temporary Exclusion Criteria: Moderate or severe acute illness/infection (according to 
investigator judgment) on the day of vaccination or febrile illness (temperature ≥ 38.0°C [≥ 
100.4°F]). A prospective subject should not be included in the study until the condition has 
resolved or the febrile event has subsided.  If the delay for the febrile illness exceeds the window 
between screening and vaccination, or if deemed necessary by the investigator, a prospective 
subject may be re-screened once the fever has resolved. 
 
6.6 
Medical History 
 
Prior to enrollment, subjects will be assessed for pre-existing conditions and illnesses, both past 
and ongoing. This assessment will include a physical examination that will include, but not be 
limited to, the following body systems: neurological, heart, head, respiratory, and abdomen. Any 
pre-existing conditions and illnesses will be documented in the source document.  Medical records 
will not be reviewed.  Significant medical history (reported as diagnosis) including conditions for 
which the subject is or has been followed by a physician or conditions that could resume during 
the course of the study or lead to an SAE or to a repetitive outpatient care will be collected in the 
electronic case report form (eCRF).  
 
Dates, medications, and body systems will be recorded; the information collected will not be coded.  
 
For each condition, the data collected will be limited to: 
 
• Diagnosis with dates (this is preferable to reporting signs and symptoms) 
 
• Presence or absence of the condition at enrollment 
 
Vitals signs will include temperature, blood pressure, and pulse taken while the subject is in a 
resting position. 
 
At subsequent visits, a history-driven physical examination will include, but not be limited to, the 
following body systems: neurological, heart, head, respiratory, and abdomen. The necessity of this 
physical examination is determined through conversation between the study staff and the subject 
about the subject’s current medical status. 
 
 
6.7 
Contraindications for Subsequent Vaccinations 
 
 
6.7.1 
Temporary Contraindications 
 
Should a subject experience one of the conditions listed below, the Investigator will postpone 
further vaccination until the condition is resolved.  
 
 
• Ongoing clinical AE or biological abnormality related to the trial vaccination 
• Febrile illness (temperature ≥ 38.0°C [≥ 100.4°F]) or moderate or severe acute 
illness/infection on the day of vaccination, according to Investigator judgment. 
Postponement must still be within the timeframe for vaccination indicated in the Tables of 
Study Procedures.  If the delay for the febrile illness exceeds the window between screening 


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and vaccination, or if deemed necessary by the investigator, a prospective subject may be 
re-screened once the fever has resolved. 
• Acute illness on day of vaccination that the investigator determines the subject to be 
ineligible for vaccine that day. 
 
6.7.2 
Definitive Contraindications 
 
Should a subject experience one of the conditions listed below, the Investigator will discontinue 
vaccination: 
 
• Pregnancy, as indicated by a positive urine test on vaccination days. 
 
• An anaphylactic or other significant allergic reaction to the previous dose of vaccine 
 
• Known or suspected congenital or acquired immunodeficiency; or receipt of 
immunosuppressive therapy such as anti-cancer chemotherapy or radiation therapy within the 
preceding 6 months; or long-term systemic corticosteroid therapy (prednisone or equivalent 
for more than 2 consecutive weeks within the past 3 months) 
 
 
• SAE related to the study vaccine following the trial vaccination 
 
• Participation in another clinical trial investigating a vaccine, drug, medical device, or medical 
procedure 
 
• Deprived of freedom by an administrative or court order, or in an emergency setting, or 
hospitalized involuntarily. Subjects will not be withdrawn due to contraindication but will 
be followed up for safety and possibly immunogenicity assessment. 
 
Note: If a subject who has already received at least 1 dose of the study vaccine, experiences a 
contraindication to further vaccination during the trial, they will continue to be followed for 
immunogenicity assessments and will not be discontinued from the trial; however, no additional 
vaccinations will be administered. 
 
6.8 
Other Conditions-for-Withdrawal beyond Definitive Contraindications 
 
Subjects will be informed that they have the right to withdraw from the trial at any time. 
A subject may be withdrawn from the study: 
• At the discretion of the Investigator or Sponsor due to safety concerns without the subject’s 
permission (withdrawal) 
• At the request of the subject (dropout) 
 
The following will result in automatic withdrawal or exclusion of a subject from the study: 
• Significant noncompliance with the protocol, based on the Investigator’s judgment 
 
The reason for a withdrawal or dropout should be clearly documented in the source documents 
and on the eCRF. 
 
 
Withdrawn subjects will not be replaced. 
 
 
 


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6.9 
Lost to Follow-up Procedures 
 
In the case of subjects who fail to return for a follow-up examination, documented reasonable 
effort (i.e., documented telephone calls) should be undertaken to locate or recall them, or at least to 
determine their health status while fully respecting their rights. These efforts should be 
documented in the source documents. 
 
6.10 
Classification of Subjects Who Discontinue the Trial 
 
For any subject who discontinues the trial prior to completion, the most serious reason for early 
termination will be checked in the eCRF. Reasons will be listed from the most serious to the least 
serious as follows: 
 
• Serious adverse event: To be used when a subject drops out of or is withdrawn from the 
study by the Investigator because of the occurrence of an SAE, as defined in Section 18. 
 
• Other adverse event: To be used when a subject drops out of or is withdrawn from the study 
by the Investigator because of the occurrence of an AE other than an SAE, as defined in 
Section 17. 
 
• Non-compliance with protocol: To be used when the Investigator withdraws a subject from 
the study because of failure to follow protocol guidelines (e.g., not attending visits, not being 
available for telephone calls, not providing blood samples). This termination category may 
also be used if it is retrospectively discovered that a subject did not fulfill the eligibility 
criteria. The Investigator will provide a comment as to the specific cause of non-compliance. 
 
• Lost to follow-up: To be used when the Investigator withdraws a subject from the study 
because of failure to establish contact, as outlined in Section 6.9. The Investigator will provide 
documentation that contact was attempted (i.e., return of unsigned certified letter receipt). 
 
• Voluntary withdrawal not due to an adverse event: To be used when a subject drops out of 
the study for any reason other than those listed above. 
 
6.10.1 Follow-up of Discontinuations 
 
The site should complete all scheduled follow-ups with any subject who has prematurely 
terminated the trial because of an SAE, other type of AE, non-compliance with the protocol, or 
loss of eligibility, including definitive contraindications. Discontinued subjects will be followed 
up to 6 months after the last vaccination. 
 
For subjects where the reason for early termination was lost to follow-up or if the subject withdrew 
informed consent and specified that they do not want to be contacted again and it is documented in 
the source document, the site will not attempt to obtain further information. 
 
 
6.10.2 Follow-up and Reporting of Pregnancies 
 
Pregnancy is an exclusion criterion for enrollment in this study, but a subject could potentially 
become pregnant during her participation. In case of pregnancy and if at least one dose of the 
study vaccines has been administered, the subject will not be discontinued from the trial and will 


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be followed for immunogenicity assessment. However, no additional vaccination will be 
administered. 
 
All pregnancy cases should be reported if they occurred during the study. Study staff must then 
maintain contact with the subject to obtain information about the outcome—i.e., details about the 
delivery and the newborn, or about pregnancy termination. 
 
Pregnancy itself is not considered an AE, but any complications during pregnancy are to be 
considered as AEs, and in some cases could be considered SAEs. Spontaneous abortions, fetal 
death, stillbirth, and congenital anomalies reported in the baby are always considered as SAEs, 
and the information should be provided using an SAE form.  
 
 7   Safety Emergency Coverage 
 
Subjects in the trial will be given contact numbers for the PI and study team members in the event of 
an emergency related to the trial.  A study investigator will be available at all times for study related 
emergencies.  If emergency health care is needed, subjects should seek care through the 911 system 
or local emergency services.  Study team contact numbers should be used to notify the study team of 
an emergency after emergent health care issues have been handled. 
 
8 Modification of the Trial and Protocol 
 
Any amendments to this trial plan and protocol must be discussed with WRAIR. If agreement is 
reached concerning the need for an amendment, the PI will be responsible for completing the 
amendment and creating a new version of the protocol will replace the earlier version. All 
substantial amendments e.g. that affect the conduct of the trial or the safety of subjects, require 
IRB approval, and must also be forwarded to regulatory authorities. 
 
An administrative amendment to a protocol is one that modifies some administrative or logistical 
aspect of the trial but does not affect its design or objectives or have an impact on the subjects’ 
safety. Regulatory authorities need only be notified about administrative changes. Sites will be 
required to submit any administrative amendments to their IRB for notification and/or approval. The 
Investigator is responsible for ensuring that changes to an approved trial, during the period for 
which IRB approval has already been given, are not initiated without IRB review and approval, 
except to eliminate apparent immediate hazards to subjects. 
 
 
   9   Vaccines Administered 
Subjects in Group 1 will receive 3 doses of Rabies vaccine administered IM at Days 0, 7,21 and 365. 
Subjects in Group 2 will receive 3 doses of Rabies vaccine administered ID at Days 0, 7,21 and IM 
at Day 365. 
Subjects in Group 3 will receive 2 doses of Rabies vaccine administered IM at Days 0,7 and 365. 
Subjects in Group 4 will receive 2 doses of Rabies vaccine administered ID at Days 0,7, and IM at Day 
365.  
Subjects in Group 5 will receive 1 dose of Human Serum Albumin administered IM at Day 0.  
Subjects in Group 6 will receive 1dose of Human Serum Albumin administered ID at Day 0.  
 


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  9.1     Identity of Trial Product: Rabies Vaccine 
 
Vaccine: RabAvert® (Rabies Vaccine) produced by Novartis Vaccines and Diagnostics GmbH 
is a sterile freeze-dried vaccine obtained by growing the fixed-virus strain Flury LEP in primary 
cultures of chicken fibroblasts.  The strain Flury LEP was obtained from American Type Culture 
Collection as the 59th egg passage.  The growth medium for propagation of the virus is a 
synthetic cell culture medium with the addition of human albumin, polygeline (processed bovine 
gelatin) and antibiotics.    The virus is inactivated with β-propiolactone, and further processed by 
zonal centrifugation in sucrose density-gradient.  The vaccine is lyophilized after addition of a 
stabilizer solution that consists of buffered polygeline and potassium glutamate.    One dose of 
reconstituted vaccine contains less than 12 mg polygeline (processed bovine gelatin), less than 
0.3 mg human serum albumin, 1 mg potassium glutamate and 0.3 mg sodium EDTA.  Small 
quantities of bovine serum are used in the cell culture process.  Bovine components originate 
only from the United States, Australia and New Zealand.  Minimal amounts of chicken protein 
may be present in the final product; ovalbumin content is less than 3 ng/dose (1 mL), based on 
ELISA.  In the final vaccine, neomycin is present at < 1 μg, chlortetracycline at < 20 ng, and 
amphotericin B at < 2 ng per dose.   RabAvert® is intended for intramuscular (IM) injection.  
The vaccine contains no preservative and should be used immediately after reconstitution with 
the supplied Sterile Diluent for RabAvert® (Water for Injection).    The potency of the final 
product is determined by the US National Institute of Health (NIH) mouse potency test using the 
US reference standard.  The potency of one dose (1.0 mL) RabAvert® is at least 2.5 IU of rabies 
antigen.   RabAvert® is a white, freeze-dried vaccine for reconstitution with the water for 
injection diluent prior to use; the reconstituted vaccine is a clear to slightly opaque, colorless 
solution.    
 
Form: 
Powder and solvent for suspension for injection 
Dose: 
1.0 ml of the reconstituted vaccine 
Route: 
ID/IM 
Batch number: 
To be determined 
  
9.1.1 Composition 
 
RabAvert® (Rabies Vaccine) produced by Novartis Vaccines and Diagnostics GmbH is a sterile 
freeze-dried vaccine obtained by growing the fixed-virus strain Flury LEP in primary cultures of 
chicken fibroblasts.  The strain Flury LEP was obtained from American Type Culture Collection as 
the 59th egg passage.  The growth medium for propagation of the virus is a synthetic cell culture 
medium with the addition of human albumin, polygeline (processed bovine gelatin) and antibiotics.    
The virus is inactivated with β-propiolactone, and further processed by zonal centrifugation in a 
sucrose density-gradient.  The vaccine is lyophilized after addition of a stabilizer solution which 
consists of buffered polygeline and potassium glutamate.     
 
   Placebo:  Human Serum Albumin 0.03%. Produced by Greer Labs, for use as a sterile diluent.  
 
     Form:                                         Diluent  
     Dose:                                          1.0 ml of diluent 
     Route:                                         IM/ID 
     Batch/ Lot number:                    To be determined 


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9.1.2 
Preparation and Administration 
The vaccine comes as powder and reconstituted for injection.  Vaccine will be administered by ID 
in the forearm or IM injection in the deltoid.   
 
9.1.3 
Dose Selection and Timing 
The dose for IM vaccinations will be 1 ml for each vaccination. The dose for ID vaccinations will 
be 0.1 ml for each vaccination. Two vaccination groups will receive vaccinations on Day 0, then 
Day 7, then Day 21. The other two vaccination groups will receive vaccinations on Day 0 and then 
Day 7. 
  
   9.1.4 
Labeling and Packaging 
 
Rabies Vaccine and Human Serum Albumin: 
 
 
 9.1.5     Product Shipment 
 
Product will be purchased through the study team, who will be responsible for coordinating 
appropriate shipping and storage. 
 
 
   9.1.6 
Product Storage 
 
The Investigator will be personally responsible for product management or will designate a staff 
member to assume this responsibility. 
 
Vaccines will be kept in the Clinical Research Unit in a secure place with restricted access. 
Vaccines will be stored in a refrigerator at a temperature ranging from +2°C to +8°C. The vaccine 
must not be frozen. The temperature must be monitored and documented for the entire time that 
the vaccine is at the trial site. In case of accidental freezing or disruption of the cold chain, vaccine 
must not be administered and must be quarantined, and the Investigator or authorized designee 
should contact the Novartis representative for further instructions. 
 
9.1.7     Product Accountability 
 
Vaccine and human serum albumin accountability will be the responsibility of the study 
coordinator, who will maintain both accountability logs. 
 
 
9.1.8     Replacement Doses 
 
This is an open-label trial. In the case that a dose needs to be replaced, another vial should be 
removed from the site’s supply. 
 
9.1.9     Disposal of Unused Products 
 
As the vaccine used in this study is an available licensed vaccine, any unused product may be used 
in an additional study conducted by the PI. Any unused human serum albumin can also be used in 
additional studies.  


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10  Blinding and Code-breaking Procedures 
 
This is an open-label trial; no code breaking procedures need to be defined. 
 
 
   11 Randomization and Allocation Procedures 
 
Each subject who meets the inclusion/exclusion criteria and signs an ICF will be assigned 
to one of the groups based on block randomization. 
 
Table 2: Randomization 
 
A statistician will prepare the randomization schedule to randomly assign subject numbers to group 
and treatment.  An appropriate blocking scheme will be used to ensure that the balance between 
treatment assignments is maintained. The biostatistician from the Center for Research and 
Evaluation will provide master randomization list to programmer with database group and label 
preparers starting with subject ID numbers 0001 through 0060.  No other study team members will 
have access to this list.  The study team will not know the randomization sequence.   
Once the screening process is complete and the subject is eligible to be enrolled in the trial, the 
subject ID number will be assigned by the clinical research coordinator or designee.  Subject 
numbers will be assigned as a 4-digit subject number (0001, 0002, 0003, etc.).  The first subject 
randomized will be 0001, etc.   
Subjects will be enrolled in six groups.  When a subject is enrolled into the study, they will be 
allocated to a particular group and complete all vaccinations and follow-ups according to the 
schedule for that group. 
Subject identification numbers will not be reassigned for any reason. 
 
12 
Treatment Compliance 
 
The following measures will ensure that the vaccine doses administered comply with those planned, 
and that any non-compliance is documented so that it can be accounted for in the data analyses: 
 
• All vaccinations will be administered by qualified trial personnel. 
 
• Study Coordinator will maintain accountability records of product delivery to the trial site, 
product inventory at the site, dose(s) given to each subject, and the disposal of unused or wasted 
doses. 
 
13 
Concomitant Medications or Other Therapies 
 
At the time of enrollment, ongoing medications including other therapies should be recorded 
in the source document as well as new medications prescribed for new medical 
conditions/AEs during trial participation. 
 
Documentation in the eCRF of concomitant medication will included all medications listed in the 
source documents. 
 
Medications will be collected in the eCRF from the day of vaccination to the end of the follow-
up period in general as they may impact the response to the vaccination and impact the 


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consistency of the information collected on concomitant medications at any vaccination. 
 
Medications pertaining to the contraindications list will be reported throughout the course of the 
trial. The information reported in the eCRF for each reported medication will be limited to: 
• Trade name 
• Indication 
 
 
• Start and stop dates 
 
Dosage and administration route will not be recorded.


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14 
Management of Samples 
 
See the Tables of Study Procedures and Section 5.2 for details of the sampling schedule. 
 
 
14.1 Sample Collection 
 
 
14.1.1 Blood Samples 
 
See the Tables of Study Procedures and Section 5.2 for details of the sampling schedule. 
Immediately prior to the blood draw, the staff member performing the procedure will verify the 
subject’s identity; will obtain the pre-printed label and the sampling stage; and will attach the label 
to the tube. Blood is to be taken from the limb opposite to the one that will be used for injection of 
the Rabies vaccine. 
 
   14.1.2 
Additional Biological Samples 
 
In the event of an AE or SAE, additional biological samples (e.g., body fluid, tissue samples) 
may be collected. 
 
 
 
 14.2 Sample Preparation 
 
An overview of the lab procedures is provided here. 
 
Antibody Assays:  
At each time point, blood samples will be obtained for measurement of neutralizing antibodies 
against rabies virus.  The blood will be allowed to clot and the serum collected by centrifugation.  
The serum will be stored frozen at -70°C or below until shipment to WRAIR or partner laboratory 
for analysis.   
CMI: 
 
Blood will be obtained for evaluation of CMI responses to vaccination. At each time point 
blood will be collected and PBMCs will be separated. The PBMCs will be stored frozen at -
70°C or below until analyzed at SUNY Upstate or shipment to WRAIR for analysis. CMI 
assays will include multiparameter flow cytometry, including intracellular cytokine staining, 
and possibly CFSE-based proliferation assays and epitope mapping depending on the results 
of initial testing. 
 
Proteomics: 
 
Blood samples will be obtained for analysis of the proteins expressed after vaccination with 
Rabies vaccine.  Whole blood will be collected and processed in the proteomics core lab at 
Upstate Medical University.   
 
 
 
 
 


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14.3 Sample Storage and Shipment 
 
During storage, serum tubes are to be kept in a freezer whose temperature is set and maintained at 
-20°C or below. The temperature will be monitored and documented on the appropriate form 
during the entire trial. If it rises above -10°C for any period of time, the Clinical Logistics 
Coordinator must be notified. See the Operating Guidelines for further details. 
 
Shipments to the laboratories will be made only after appropriate monitoring, and following 
notification of the Clinical Logistics Coordinator. Sera will be shipped frozen, using dry ice to 
maintain them in a frozen state, in the packaging container provided by the carrier. Again, 
temperatures will be monitored. Shipments must be compliant with the International Air 
Transport Association (IATA) 602 regulations. 
 
Samples will be shipped to the Walter Reed Army Institute of Research and Kansas State University.  
 
14.4 Future Use of Stored Serum Samples for Research 
 
Any unused part of the research samples will be securely stored at WRAIR, or Upstate Medical 
University for at least 5 years.  
 
Subjects will be asked to indicate in the ICF whether they will permit the future use of any unused 
samples for other tests. If they refuse permission, the samples will not be used for any testing other 
than that directly related to this study. If they agree to this use, they will not be paid for giving 
permission. (Anonymity of samples will be ensured.) The aim of any possible future research is 
unknown today, and may not be related to this particular study. It may be to improve the 
knowledge of vaccines or infectious diseases, or to improve laboratory methods. Genetic tests will 
never be performed on these samples without individual informed consent. 
 
  15 Clinical Supplies 
The Investigator will supply all phlebotomy (with the exception of Vacutainers ®) and 
centrifugation equipment, including biohazard and/or safety supplies, blood collection tubes, 
cryotubes, cryotube storage boxes, cryotube labels, shipping containers. The biohazard and safety 
supplies include examination gloves, laboratory coats, and sharps disposal containers. The site will 
ensure that all biohazard wastes are disposed of in accordance with local practices. The Investigator 
will also supply appropriate space in a temperature-monitored refrigerator for the storage of the 
products and for the blood samples, and appropriate space in a temperature monitored freezer for 
serum and PBMC aliquots. 
 
 
16 
Endpoints and Assessment Methods 
16.1 
Primary Endpoints 
 
 
16.1.1 
Immunogenicity Endpoints 
 
The primary endpoints for the evaluation of immunogenicity are: 
 
1) Percentage of subjects in each group with titer of IgG antibodies above the protective limits 
( > or = 0.5 IU/ml) at 1 and 12 months after initial vaccine dose.  


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16.2      Secondary Endpoints 
 
 
   16.2.1   Immunogenicity Endpoints 
 
 
1) Total IgG and neutralizing Ab levels, measured by ELISA and PRNT, against rabies virus at 
baseline and at 28 days 1 month and 365 days post-initial vaccine dose. 
 
16.3  Assessment Methods 
 
 
16.3.1     Immediate Post-vaccination Surveillance Period 
 
Subjects will be kept under observation for 30 minutes after each Rabies vaccination to 
ensure their safety. Any AE that occurs during this period will be noted on the source 
document and identified as an immediate event / reaction; and will additionally be recorded in 
the eCRF. 
 


 
 
 
17 
Reporting AEs to IRBs 
 
 
The Investigators will be responsible for informing the IRBs that reviewed the trial protocol 
of any reportable SAEs. 
 
  18 Reporting of Serious Adverse Events 
 
In order to comply with current regulations on SAE reporting to health authorities, the 
Investigator must document all SAEs regardless of causal relationship, and notify the 
Sponsor within the notification timelines stated in the following sections. The Investigator 
will give access and provide the Sponsor and the local clinical research associate (CRA) with 
all necessary information to allow the Sponsor to conduct a detailed analysis of the safety of 
the investigational product. It is the responsibility of the Investigator to request all 
necessary documentation (e.g., medical records, discharge summary, autopsy) in order to 
provide comprehensive safety information. All relevant information must then be 
transcribed into the SAE form.   
 
  Timelines and process for SAE reporting    
Initial reports of SAEs should be sent by the Upstate PI within 24 hours of the Investigator 
becoming aware of an SAE.  In parallel, the Medical Monitor for the study should be 
notified. The Human Reasearch Protection Office (HRPO) should also be notified via email: 
usarmy.detrick.medcom-usamrmc.other.hrpo@mail.mil  Human Research Protection Office 
(HRPO), Office of Research Protections (ORP), U.S. Army Medical Research and Materiel 
Command, 810 Schreider Street 
Frederick, MD 21702-5012. 
 
Process for SAE documentation   
Following receipt of SAE reports, the Sponsor may raise additional questions to ensure 
complete documentation of the SAEs.  
Timelines and process for pregnancy reporting  
If the Investigator becomes aware on pregnancy of study participant, paper pregnancy 
reporting form should be completed. The Investigator will notify the medical monitor email: 
fazilit@upstate.edu and HRPO email: usarmy.detrick.medcom-
usamrmc.other.hrpo@mail.mil  Human Research Protection Office (HRPO), Office of 
Research Protections (ORP), U.S. Army Medical Research and Materiel Command, 810 
Schreider Street 
Frederick, MD 21702-5012. 
 
18.1 Follow-up Reporting by the Investigator 
 


 
 
The SAE Form completed initially must be updated within 24 hours after the 
Investigator has become aware of any new relevant information concerning the SAE 
(e.g., outcome, precise description of medical history, results of the investigation). 
Copies of documents (e.g., medical records, discharge summary, autopsy) may be 
requested  
 
 
The anonymity of the subject must always be respected when forwarding this information. 
 
 
  18.2 Reporting of SAEs Occurring After a Subject Has Completed the 
Study 
 
Any SAE that occurs after a subject has completed the study but that is likely to be 
related to the product or to the experiment must also be reported as soon as possible.  
 
18.3 Assessment of Causality 
 
The causal relationship between the SAE and the product will first be evaluated by the 
Investigator, using the following definitions: 
 
0 - Not related: The AE is clearly / most probably caused by other etiologies such as 
an underlying condition, therapeutic intervention, or concomitant therapy; or the 
delay between vaccination and the onset of the SAE is incompatible with a causal 
relationship; or the SAE started before the first vaccination. 
 
1 - Related: There is a “reasonable possibility” that the SAE was caused by the 
vaccination, meaning that there is evidence or arguments to suggest a causal 
relationship. 
 
(ICH Guidelines, Clinical Safety Data Management E2A) 
 
Following this, the Sponsor’s Product Safety Officer (PSO) will also assess the causal 
relationship to the product, based on the available information and current medical 
knowledge. 
 
The decision to modify or discontinue the trial may be made after mutual agreement 
between the Sponsor and the Investigators. 
 
   
18.4 
Reporting SAEs to Health Authorities and IRBs 
 
The Sponsor will inform the relevant health authorities of any reportable SAEs 
according to the local regulatory requirements. Reporting to the health authorities will 
be done according to the Sponsor’s SOPs. 
 
The Sponsor’s Responsible Medical Officer will notify the Investigators in writing of the 
occurrence of any reportable SAEs. The Investigators will be responsible for informing 
the IRBs that reviewed the trial protocol of any reportable SAEs. 
 
Per requirements of Department of Defense Instructions 3216.02, research funded with 


 
 
Department of Defense funds requires a research monitor to be appointed.  Dr. Tasaduq 
Fazili will be the research monitor for this trial.  His duties will include reviewing and 
providing and unbiased assessment and written report of all: SAEs, Subject deaths, 
Unexpected AEs, Protocol deviations that are related to an adverse event impacting the 
subject, Annual reports and Unanticipated problems involving risks to subject and others 
involved in the trial.  The research monitor may observe recruitment and enrollment 
procedures and the consent process for individuals, groups or units, review monitoring 
plans and data collection. 
The research monitor may discuss the research protocol with the investigators, interview 
human subjects, and consult with others outside of the study about the research.  He shall 
have authority to stop a research protocol in progress, remove individual human subjects 
from a research protocol, and take whatever steps are necessary to protect the safety and 
well-being of human subjects until the IRB can assess the monitor's report.  He shall have 
the responsibility to promptly report his observations and findings to the IRB or other 
designated official and the Human Research Protection Office. 
 
19 
Data 
 
19.1 Data Collection and eCRF Completion 
 
At specified intervals, the Investigator or an authorized designee will interview the 
subjects to collect information, and will attempt to clarify anything that is incomplete or 
unclear. All clinical trial information gathered by the study site will be reported 
electronically by the Investigator or authorized designee using a web-based eCRF. The 
eCRF has been designed specifically for this trial using a validated Electronic Records / 
Electronic Signature-compliant platform (21CFR Part 11).   
Upon completion of training, each user requiring access to the EDC system will be 
issued a unique username and password. In the event of a change in trial personnel, each 
newly assigned individual will receive a unique username and password; the username 
and password of a previous user may not be reissued.  
The Investigator is responsible for the timeliness, completeness, and accuracy of the 
information in the eCRFs; must provide explanations for all missing information; and 
must sign the eCRF using an e-signature. 
 
 
19.2 Data Management 
 
Management of Clinical Data 
 
Data generated during the trial will be managed following two different processes: 
• Clinical data, defined as all data reported in the eCRF, and laboratory data will be 
entered in the database held at Frontier Science in Amherst, NY. 
 
During the trial, clinical data reported in the eCRFs will be integrated into the clinical 


 
 
database under the responsibility of Upstate.  Data monitoring at the sites and quality 
control in the form of computerized logic and / or consistency checks will be 
systematically applied in order to detect errors or omissions. Any questions pertaining 
to the reported clinical data will be submitted to the Investigator for resolution using 
the EDC system. Each step of this process will be monitored through the 
implementation of individual passwords to maintain appropriate database access and to 
ensure database integrity. 
 
The validation of the immunogenicity data will be performed at the laboratory level 
following the laboratory’s procedures. Information from the laboratory will be checked 
for consistency before integration into the clinical database. 
 
After integration of all corrections in the complete set of data, the database will be released 
for statistical analysis. 
 
19.3 Data Review 
 
A review of the data is anticipated through the data review process led by Data Management 
before database lock. 
 
 
20 
Statistical Methods 
 
The analysis will be performed under the responsibility of Upstate Medical University 
in conjunction with the study partners. 
 
This will be a descriptive study to assess immunogenicity of a licensed rabies vaccine using 
test dosage schedules and routes of administration. No formal statistical hypothesis tests will 
be conducted.. Descriptive analyses for the primary endpoints will be based on per-protocol 
analysis data sets using the data from blood samples taken at baseline (0 days), 28 days, and 
365 days after the initial dose and at 7 days after a booster at one year (372 days).. Separate 
descriptive analyses will be conducted for each per-protocol data set and group. 
 
Exploratory statistical analyses of final data may be conducted, if indicated by the 
descriptive results. Parametric, non-parametric and resampling (bootstrapping) methods for 
statistical inference may be used in exploratory analyses, based on data compliance with 
assumptions of methods.  P -values  ≤ 0.05 will be considered significant and p-values ≤ 
0.10 will be considered a trend. Confidence intervals will be constructed at α=0.05 and 
α=0.10.  When necessary, p-value corrections for multiple comparisons will be applied. 
 
A descriptive summary of adverse events that were reported throughout the trial will be 
produced. Descriptive and inferential analyses (if any) will be carried out using SAS Version 
9.2 (or more recent versions), which is licensed and supplied by SAS Institute, Cary, NC, 
USA. 
 
20.1 
Statistical Methods for Primary Immunogenicity Objectives 
 
Immunogenicity against rabies will be assessed descriptively using the following 


 
 
parameters: 
Timepoints:  At baseline, 28 days and 372 days.  
 
 • Number and percentage of subjects with a titer ≥ 0.5 IU/ml against rabies virus by 
treatment group at days 28 and 372. 
 
 
The descriptive statistics listed above will be calculated for each of the six groups for each 
blood draw.  Descriptive results will be reported in tabular and graphic formats.  A separate 
document, the statistical analysis plan (SAP) will be developed prior to data analysis.  It will 
provide more detailed information about the specific descriptive statistics to be used and the 
format (tables, listings and figures) by which the statistics will be reported. 
 
 
20.2 
Statistical Methods for Secondary Immunogenicity Objectives 
 
Immunogenicity will be assessed descriptively using the following parameters: 
 
Timepoints:  Day 28 and day 365.  
 
The descriptive statistics listed above will be calculated for each of the six groups for the 
time points stated above.  Specification of statistics and reporting formats will be provided in 
the SAP, as described above. 
 
20.3 
Per-Protocol Analysis Set 
 
The Per-Protocol (PP) analysis set will include all subjects who had no protocol 
deviations at the time that the data was collected. Three PP analysis sets will be 
produced for the primary objective that will contain separate data from blood 
drawn on day 0, day 28 and day 372, respectively. Descriptive analysis of the 
secondary endpoint will require a fourth per protocol analysis set for blood drawn 
on day 365.  Subjects will be excluded from the PPAS for the following reasons: 
  1)  Subject did not meet all protocol-specified inclusion/exclusion criteria 
or   definitive contraindications (only for the second and third 
vaccination). 
 
 
 2)  Subject did not receive the correct number of injections. 
 
 
 
 3)  Administration of vaccine was not done as per protocol (site and route of 
administration). 
 
   4)  Subject did not receive vaccine in the proper time window defined in the tables 
of the         study procedures. 
 
 5)  Subject did not provide a post-dose serology sample in the proper time window 
defined in  
         the tables of the study procedures. 


 
 
 
Subjects will remain in this population as long as they do not meet one of the above criteria, 
except for blood sampling timing and validity of the serology test result*. A PP set will be 
defined for each injection. 
 
*Note: For example, a subject whose time period is outside the defined visit window 
only between the first injection and the blood sample taken after the first injection will 
be excluded from PP1 but may be kept in PP2 or PP3. 
 
Subjects will be analyzed by the vaccine group to which they were randomized, subject to 
the exclusion criteria above 
 
  20.4 
Full Analysis Set 
 
The full analysis set (FAS) is defined as the subjects who received at least one injection of 
Rabies vaccine and had at least one blood sample drawn and valid post-injection serology 
result (i.e. a result different from “NR” or missing). 
 
Subjects will be analyzed by the treatment group to which they were randomized. 
 
20.5 
Populations Used in Analyses 
 
The immunogenicity analysis will be performed on the PP.  The kinetic analysis will be 
performed       on the FAS. 
 
 
 
21 
Handling of Missing Data and Outliers 
 
 
21.1 
Immunogenicity 
 
For the computation of GMTs, a titer reported as < LLOQ will be converted to a value of 
0.5 LLOQ. 
 
For calculating fold-rise and titer ratio (GMTR), < LLOQ will be converted to 0.5 
LLOQ for a numerator and < LLOQ will be converted to LLOQ for a denominator 
 
Any titer reported as > ULOQ (upper limit of quantization) will be converted to 
ULOQ.  
 
Missing data will not be imputed.  
 
Potential outliers will be identified using histograms and modified box plots or normal 
probability plots, when appropriate.  Choice of subsequent tests for outliers (e.g. Grubbs, 
Tietjen-Moore and ESD) will depend on the number and nature of suspected outliers and 
may require data transformation. Bootstrapped estimates of standard errors may also be 
reported. Sensitivity analysis of effect of outliers on descriptive statistics will be 
conducted, and if necessary descriptive statistics that include and exclude outliers will be 
reported.  
 
 22 
Determination of Sample Size and Power Calculation 


 
 
 
The planned sample size is 60 subjects, who will be randomly allocated (2:2:2:2:1:1) to six 
treatment groups.  The sample size was chosen empirically, based on sample sizes of other 
exploratory studies of this general type.  This exploratory study is not powered to formally 
test equivalency or non-inferiority in immunogenicity among groups according to 
vaccination dosage and route of administration. 
 
 
23  Ethical and Legal Issues and Investigator / Sponsor 
Responsibilities 
 
23.1 Ethical Conduct of the Trial / Good Clinical Practice 
 
This trial will be conducted in accordance with the Seoul revision of the Declaration of 
Helsinki as far as adopted by the concerned regulatory authorities, as well as in 
accordance with GCP as defined by International Conference on Harmonization (ICH), 
and with the applicable national and local requirements regarding ethical committee 
review, informed consent, and other statutes 
or regulations regarding the protection of the rights and welfare of human subjects 
participating in biomedical research. 
 
23.2 Source Data and Source Documents 
 
“Source data” are the data contained in source documents. Source documents are original 
documents or certified copies, and include, but are not limited to medical and hospital 
records, screening logs, informed consent / assent forms, Telephone Contact Logs, and 
worksheets. The purpose of trial source documents is to document the existence of 
subjects and to substantiate the integrity of the trial data collected. Investigators must 
maintain source documents so that they are accurate, complete, legible, and up to date. 
 
The subject screening and enrollment log should list all individuals contacted by the 
Investigators to participate in the trial, regardless of the outcome. 
 
 
23.3 Confidentiality of Data and Access to Subject Records 
 
Prior to initiation of the trial, the Investigator will sign a fully executed confidentiality 
agreement. 
 
The IRBs, and regulatory agencies, including the FDA, require direct access to all study 
records, and will treat these documents in a confidential manner. 
  
 
 
23.4 Monitoring, Auditing, and Archiving 
 
23.4.1 Monitoring 
 
Before the start of the trial (i.e., before the inclusion of the first subject), the Investigators 
and the Monitoring representative will meet at the site-initiation visit to discuss the trial 


 
 
protocol and the detailed trial procedures. Emphasis will be placed on inclusion and 
exclusion criteria, visit timing, informed consent procedures, eCRF completion, and the 
handling of samples and products. 
 
The monitoring staff will ensure and document that all material to be used during the trial 
has been received at the site; and that the study investigator team and local monitoring staff 
have been properly informed about the trial, GCP and regulatory requirements. 
 
After the start of the trial, the monitoring staff or a representative will be in regular 
contact with the investigational team through telephone calls and regular follow-up 
visits. The Investigator or delegate must be available for these visits, and must allow the 
monitoring staff direct access to subject medical files and eCRFs. During these visits, 
the monitoring staff will: 
 
• Evaluate the quality of the trial progress (adherence to protocol and any study-specific 
guidelines, quality of data collection and document completion, signature of consent 
forms, occurrence of SAEs, sample and product management, cold-chain monitoring, 
archiving, and direct observation of study procedures during the trial.) 
 
• Source-verify completed eCRFs and any corresponding answered queries 
 
• Determine the number of complete or ongoing issues identified at monitoring visits 
(e.g., protocol violations, SAEs). Any identified problems will be discussed with the 
Investigator, and corrective or preventive actions will be determined, as appropriate. 
 
•  review that all protocol procedures have been completed and the data have been 
entered into the eCRF. All data-related queries must be completed prior to database 
lock. 
 
At the end of the trial, a close-out visit will be performed to ensure that: 
 
• The center has all the documents necessary for archiving. 
 
• All samples have been shipped to the appropriate laboratories. 
 
• All unused materials and products have been either destroyed or returned. 
 
23.4.2 Audits and Inspections 
 
A quality assurance (QA) audit may be performed at any time to verify that the trial has 
been conducted according to the protocol, GCP and ICH requirements, and other 
applicable regulations. An inspection may be conducted by regulatory authorities. The 
Investigator must allow direct access to trial documents during these inspections and 
audits. 
 
23.4.3 Archiving 
 
The Investigator will retain all source documents, consent forms, for 2 years after the 
investigation is discontinued. 
The Investigator will retain all documentation pertaining to the trial. Archived data may 
be held on microfiche or electronic records, provided that a back-up exists and that a 
hard copy can be obtained if required. The protocol, documentation, approvals, and all 
other documents related to the trial, including certificates attesting that satisfactory audit 
and inspection procedures have been carried out will be kept by Upstate in the 


 
 
Investigator’s File. Data on AEs are included in the Investigator’s file. All data and 
documents will be made available if requested by relevant authorities. 
 
 
24 
Stipends for Participation 
 
Subjects will receive compensation for each study visit.  Specifics of compensation 
will be included in the ICF and approved by the Upstate IRB. 
 
 
25 
 Reference List 
 
 
1. 
Plotkin SA. Rabies. Clin Infect Dis. [Review]. 2000 Jan;30(1):4-12. 
2. 
Briggs D, Hanlon CA. World Rabies Day: focusing attention on a neglected disease. 
Vet Rec. [Review]. 2007 Sep 1;161(9):288-9. 
3. 
Strady A, Lang J, Lienard M, Blondeau C, Jaussaud R, Plotkin SA. Antibody 
persistence following preexposure regimens of cell-culture rabies vaccines: 10-year 
follow-up and proposal for a new booster policy. J Infect Dis. [Clinical Trial 
Comparative Study, Controlled Clinical Trial, Research Support, Non-U.S. Gov't]. 1998 
May;177(5):1290-5. 
4. 
Shantavasinkul P, Tantawichien T, Jaijaroensup W, Lertjarutorn S, Banjongkasaena A, 
Wilde H, et al. A 4-site, single-visit intradermal postexposure prophylaxis regimen for 
previously vaccinated patients: experiences with >5000 patients. Clin Infect Dis. 
[Research Support, Non-U.S. Gov't]. 2010 Nov 1;51(9):1070-2. 
5. 
Feyssaguet M, Dacheux L, Audry L, Compoint A, Morize JL, Blanchard I, et al. 
Multicenter comparative study of a new ELISA, PLATELIA RABIES II, for the 
detection and titration of anti-rabies glycoprotein antibodies and comparison with the 
rapid fluorescent focus inhibition test (RFFIT) on human samples from vaccinated and 
non-vaccinated people. Vaccine. [Comparative Study, Multicenter Study]. 2007 Mar 
8;25(12):2244-51. 
6. 
Welch RJ, Anderson BL, Litwin CM. An evaluation of two commercially available 
ELISAs and one in-house reference laboratory ELISA for the determination of human 
anti-rabies virus antibodies. J Med Microbiol. [Comparative Study, Evaluation Studies, 
Research Support, Non-U.S. Gov't]. 2009 Jun;58(Pt 6):806-10. 
7. 
Aebersold R, Mann M. Mass spectrometry-based proteomics. Nature. [Research 
Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. Review]. 2003 Mar 
13;422(6928):198-207. 
8. 
Mann M, Jensen ON. Proteomic analysis of post-translational modifications. Nat 
Biotechnol. 2003 Mar;21(3):255-61. 
9. 
Shartava A, Monteiro CA, Bencsath FA, Schneider K, Chait BT, Gussio R, et al. A 
posttranslational modification of beta-actin contributes to the slow dissociation of the 
spectrin-protein 4.1-actin complex of irreversibly sickled cells. J Cell Biol. 1995 
Mar;128(5):805-18. 
10. Cong YS, Fan E, Wang E. Simultaneous proteomic profiling of four different growth 


 
 
states of human fibroblasts, using amine-reactive isobaric tagging reagents and tandem 
mass spectrometry. Mech Ageing Dev. 2006 Apr;127(4):332-43.  
11.   Rupprecht CE, Plotkin SA: Chapter 29: Rabies Vaccines in Vaccines, 6th edition, edited 
by Plotkin SA, Orenstein WA, Offit PA.  Elsevier Inc., 2013: 663. 

SAP_001.pdf: 
Title: Immunogenicity of a Two vs Three dose, Intradermal (ID) vs Intramuscular (IM) Administration of a 
Licensed Rabies Vaccine for Pre-Exposure Vaccination 
NCT number: 02374814 
Document date: 05 December 2016 
 
 


Immunogenicity of a Two vs Three dose, Intradermal (ID) vs Intramuscular (IM) 
Administration of a Licensed Rabies Vaccine for Pre-Exposure Vaccination 
 
Randomized, open-label, single center trial in adults aged 18 to 60 years in the US 
 
 
 
 
Statistical Analysis Plan (SAP) 
Version 1.1 Dated 05 Dec 2016 
 
 
 
 
Funder’s Representative 
LTC(P) Paul Keiser 
 
Walter Reed Army Institute of Research 
 
503 Robert Grant Avenue 
 
Silver Spring, MD 20910 
 
 Principle Investigator: 
Mark E Polhemus 
 
Upstate Medical University 
 
725 Irving Ave. Suite 311 
 
Syracuse, NY 13210 
 
Licensed Products:                                    The RabAvert® Rabies Vaccine                                         
 
Form /Route: 
 Intradermal and Intramusclar 
 
Indication For This Study: 
 Comparison of Dose and Schedule   
                
 
Medical                                                       Donald Blair, MD 
Officer: 
Upstate Medical University  
 
 
 
 
 
 
 
Funder’s Clinical Development  
Lou Jasper 
Director: 
USAMMDA 
 
1430 Veteran’s Drive 
 
Ft. Detrick, MD 21702 
 
Tel: (301) 619-4951  
 
Fax: (301) 619-2304 
 
 
Study Site/PI Location: 
Center for Global Health and Translational Science 
 
Upstate Medical University 
 
Syracuse, NY 
 
Study Statistician and 
Donald A. Cibula, PhD 
SAP Author: 
Associate Professor, Department of Health and Preventive Medicine 
 
SUNY Upstate Medical University 
 
2263 Weiskotten Hall 
 
Syracuse, NY 13210 
 
 


Statistical Methods 
The analysis will be performed under the responsibility of Upstate Medical University in conjunction 
with the study partners. 
This will be a descriptive study to assess immunogenicity of a licensed rabies vaccine using test dosage 
schedules and routes of administration. No formal statistical hypothesis tests will be conducted 
Descriptive analyses for the primary endpoints will be based on per-protocol analysis data sets using the 
data from blood samples taken at baseline (0 days), 28 days, and 365 days after the initial dose and at 7 
days after a booster at one year (372 days). Separate descriptive analyses will be conducted for each 
per-protocol data set and group. 
Exploratory statistical analyses of final data may be conducted, if indicated by the descriptive results. 
Parametric, non-parametric and resampling (bootstrapping) methods for statistical inference may be 
used in exploratory analyses, based on data compliance with assumptions of methods.  P -values ≤ 0.05 
will be considered significant and p-values ≤ 0.10 will be considered a trend. Confidence intervals will be 
constructed at α=0.05 and α=0.10.  When necessary, p-value corrections for multiple comparisons will 
be applied. 
A descriptive summary of adverse events that were reported throughout the trial will be produced. 
Descriptive and inferential analyses (if any) will be carried out using SAS Version 9.2 (or more recent 
versions), which is licensed and supplied by SAS Institute, Cary, NC, USA. 
Per-Protocol Analysis Set 
The Per-Protocol (PP) analysis set will include all subjects who had no protocol deviations at the time 
that the data was collected. Three PP analysis sets will be produced for the primary objective that will 
contain separate data from blood drawn on day 0, day 28 and day 372, respectively. Descriptive analysis 
of the secondary endpoint will require a fourth per protocol analysis set for blood drawn on day 365.  
Subjects will be excluded from the PPAS for the following reasons: 
1. Subject did not meet all protocol-specified inclusion/exclusion criteria or definitive 
contraindications (only for the second and third vaccination) 
2. Subject did not receive the correct number of injections 
3. Administration of vaccine was not done as per protocol (site and route of administration) 
4. Subject did not receive vaccine in the proper time window defined in the tables of the study 
procedures 
5. Subject did not provide a post-dose serology sample in the proper time window defined in the 
tables of the study procedures 
Subjects will remain in this population as long as they do not meet one of the above criteria, except for 
blood sampling timing and validity of the serology test result*. A PP set will be defined for each 
injection. 


*Note: For example, a subject whose time period is outside the defined visit window only between the 
first injection and the blood sample taken after the first injection will be excluded from PP1 but may be 
kept in PP2 or PP3. 
Subjects will be analyzed by the vaccine group to which they were randomized, subject to the exclusion 
criteria above 
Full Analysis Set 
The full analysis set (FAS) is defined as the subjects who received at least one injection of Rabies vaccine 
and had at least one blood sample drawn and valid post-injection serology result (i.e. a result different 
from “NR” or missing). 
Subjects will be analyzed by the treatment group to which they were randomized. 
Populations Used in Analyses 
The immunogenicity analysis will be performed on the PP.  The kinetic analysis will be performed on the 
FAS. 
Handling of Missing Data and Outliers 
Immunogenicity 
For the computation of GMTs, a titer reported as < LLOQ will be converted to a value of 0.5 LLOQ. 
For calculating fold-rise and titer ratio (GMTR), < LLOQ will be converted to 0.5 LLOQ for a numerator 
and < LLOQ will be converted to LLOQ for a denominator 
Any titer reported as > ULOQ (upper limit of quantization) will be converted to ULOQ.  
Missing data will not be imputed.  
Potential outliers will be identified using histograms and modified box plots or normal probability plots, 
when appropriate.  Choice of subsequent tests for outliers (e.g. Grubbs, Tietjen-Moore and ESD) will 
depend on the number and nature of suspected outliers and may require data transformation.  .  
Bootstrapped estimates of standard errors may also be reported.  Sensitivity analysis of effect of outliers 
on descriptive statistics will be conducted, and if necessary descriptive statistics that include and 
exclude outliers will be reported 
Statistical Methods for Baseline Comparisons 
Mean (SD) and median (IQR) age at baseline will be computed and descriptively compared across the six 
groups.  Results will be presented in tabular format. 
The number and percentage of males and females per group at baseline will be calculated and results 
will be presented in tabular format. 


Baseline mean (SD) and median (IQR) of all other quantitative demographic variables of interest will be 
calculated for each group and the results will be presented in a table. 
The number and percentage per group at baseline of all other qualitative demographic variables of 
interest will be calculated for each group and the results will be presented in tabular format. 
Statistical Methods for Subject Disposition 
For each group, the number and percentage of subjects who received a total of 0, 1, 2, 3 and 4 rabies 
vaccinations will be reported in tabular format. 
Statistical Methods for Primary Immunogenicity Objectives 
Immunogenicity against rabies will be assessed descriptively using the following parameters: 
Timepoints:  At baseline, 28 days and 372 days.  
• 
Number and percentage of subjects with a titer ≥ 0.5 IU/ml against rabies virus by treatment 
group at days 0, 28 and 372. 
A table that presents the total number of per-protocol subjects and the number and percent of each 
treatment group who have a titer ≥ 0.5 IU/ml against rabies virus at days 0, 28 and 372 will be produced.  
Statistical Methods for Secondary Immunogenicity Objectives 
Immunogenicity will be assessed descriptively using the following parameters: 
Timepoints: Days 0, 28 and 365.  
A table that presents, by treatment group, the total number of per protocol subjects and the number 
and percent of each group who have IgG titers ≥ 0.5 IU/ml against rabies virus at days 0, 28 and 365 will 
be produced. 
Statistical Methods for Adverse Events and Serious Adverse Events 
For each treatment group, a line listing by subject that describes adverse events and post-baseline day 
of occurrence of the AE will be produced.   
The number and percent of subjects in each group who reported any AE between Days 0 and 372 will be 
summarized in tabular format. 
A table showing the number and percentage of subjects in each group who reported adverse events by 
general type (category) of event will be produced.