A Phase I/II Study Of The Immunogenicity Of The Yellow Fever Vaccine 17D (YFVax®) In Adults With Prior 17D Vaccination
Phase: PHASE1
Conditions: Yellow Fever, Immunization; Infection
Sponsor: Oregon Health and Science University

 
1 
Section 
Version 2.0 changes 
 
 
 
 
Clinical 
Protocol  
Replace Version 1.0 with Version 3.0 
Update protocol date 
 
 
 
 
 

 
2 
CLINICAL PROTOCOL 
 
PROTOCOL TITLE: A phase I/II study of the immunogenicity of the yellow fever vaccine 17D 
(YFVax®) in adults with prior 17D vaccination. 
DATE: 21 APRIL 2023 
PHASE: 
I/II  
IND SPONSOR: 
Food and Drug Administration 
Silver Spring, MD 200993-002 
Principle investigator  William Messer, MD PhD 
Oregon Health & Science University  
Tel: (503) 494-2185 
Email:messer@ohsu.edu 
 
Trial Identifier Pending 
Version 
3.0 
  
 
 

 
3 
1 
SYNOPSIS 
 
Title of the Protocol: A phase I/II study of the immunogenicity of the yellow fever vaccine 
17D (YFVax®) in adults with prior 17D vaccination. 
 
Overview: Yellow fever virus (YFV) is the prototype flavivirus, closely related to dengue (DENV), 
Zika (ZIKV) and Japanese encephalitis (JEV) viruses. YFV is transmitted primarily by Aedes 
aegypti mosquitoes in urban transmission cycles and by Haemogogus spp. forest canopy 
mosquitoes in sylvatic, non-human primate transmission cycles. Historically YFV has been the 
most important arthropod-borne viral pathogen of humans worldwide, causing explosive 
epidemics in Europe, the Americas and Africa in the 18th, 19th and early 20th centuries, and it 
remains endemic to at least 34 African and 13 South American countries. Control of YFV has 
been accomplished largely through vector abatement and mass vaccination with the YFV 
vaccine 17D. In the early 2000’s the WHO estimated that there were approximately 200,000 
cases/year, with 30,000 deaths, but in the past decade falling vaccination rates and spillover of 
sylvatic YFV have led to a dramatic resurgence of YFV disease, with an estimated 130,000 
severe cases and 78,000 fatalities in Africa in 2013, an estimated 7,334 sylvatic-urban cases in 
Angola and the Democratic Republic of Congo in 2015-2016, and a series of sylvatic outbreaks 
in Brazil since 2016 that threaten to spill into urban settings.  
Since its development by Max Theiler in 1937, the live-attenuated virus (LAV) YFV vaccine 
17D—administered via population-wide vaccination—has been the cornerstone of YFV control. 
Despite being widely regarded as highly immunogenic and durable, historically the vaccine has 
been administered as a prime followed by boosting every 10 years. However, the rise in global 
demand for vaccine doses because of recent YFV outbreaks has resulted in unprecedented 
worldwide shortages of 17D. These critical shortages have led to strategies to extend vaccine 
stocks, including a directive by the CDC and WHO to replace the long-standing 10-year boost 
with once-in-a-lifetime vaccination for most vaccinees. The policy changes should relieve short-
term pressure on the vaccine supply, but they move 17D vaccination policies into uncharted 
waters. Scrutiny and criticism of these strategies, particularly that of dropping the 10-year boost, 
make clear that definitive studies to validate or revise these strategies have not been done. 
Specifically, the immune correlates of protection, how the vaccine induces protective immunity, 
and who maintains and who loses long-term protection have not been well-defined.  
 
Rationale: In 1973 Mason et al demonstrated a clear correlation between 17D induced 
neutralizing antibody titers and protection against lethal challenge with the virulent YFV Asibi 
strain in Rhesus macaques. Subsequently, neutralizing antibody titers have become the most 
widely accepted correlate of 17D-induced protection and are believed to be necessary and 
sufficient for YFV immunity. Generalizability of Mason’s data is difficult, however, because 
Mason’s protective threshold titer was demonstrated in hon-human primates and based on a 
ratio between log10 pre- and post-vaccination neutralizing antibody titers – the log neutralizing 
index (LNI) –equivalent LNI or alternate protective neutralizing antibody titer has never been 
validated in humans. Nevertheless, 90% percent plaque reduction neutralization test (PRNT90 
or NT90) titers >1:10 are often taken as an immune correlate of protection, despite a lack of 
supporting data. A limited number of studies using 17D booster doses as a live-attenuated 
challenge model (boosting is a true homologous live-virus challenge) have evaluated 
neutralized antibody titers that confer “sterilizing” immunity. In these studies, sterilizing immunity 
is defined as a less than four-fold rise in neutralizing antibody titer post-boost and/or no 
detection of vaccine virus by virus isolation or PCR.  These studies suggest that NT80 or NT90 
titers >1:40 are probably required for reliable sterilizing immunity against live attenuated 17D 
challenge. Drawing upon the model that 17D boost is a homologous live-virus challenge, we will 
prospectively recruit and boost 35 individuals with history of 1 prior 17D vaccination ~10 years 

 
4 
Title of the Protocol: A phase I/II study of the immunogenicity of the yellow fever vaccine 
17D (YFVax®) in adults with prior 17D vaccination. 
 
prior. We will characterize pre- and post-boost neutralizing antibody titers, boost vaccine viral 
load, and innate and B-cell responses in boosted vaccinees.  We will identify titers below which 
vaccine viral load and/or antibody boost responses are detected and higher titers that confer 
sterilizing immunity without detectable viral load or antibody titer boost, establishing both 
stringent and relaxed correlates of antibody protection. We will also assess whether some 
vaccinees with undetectable neutralizing antibodies are still protected against 17D challenge, 
setting the stage to explore the extent to which protection may be provided by other immune 
mechanisms. 
 
Study Design: This is a prospective single arm study of the immunogenicity of the yellow fever 
vaccine 17D (YFVax®, Sanofi Pasteur, Swiftwater, PA) in adults who have documented history 
of prior yellow fever vaccination 8 or more years prior. Study subjects will receive a single YFV-
17D vaccination and be followed for 30 days after vaccination with blood samples collected on 
days 1, 2, 4, 6, 8, 10, 12, 14 and 28 post vaccination. The study is designed to enroll 34 
vaccinees. Study duration is expected to be no longer than 60 days.  Subject accrual will be 
completed in 2 years.   
  
Objectives: 
Primary Objective: To assess 17D vaccine viremia and antibody titer boost post vaccination. 
Secondary Objectives: To assess the magnitude and specific innate immune responses to 
vaccination, to asses pre- and post-vaccination B and T-cell populations and responses to 
vaccination. 
Safety evaluations: Diary cards will record solicited adverse events (AEs) for local reactions for 
7 days following vaccination and solicited systemic AEs for 14 days post vaccination. 
Unsolicited AEs will be monitored for 28 days after vaccination. 
 
Study Population: 
Inclusion Criteria: 
Male or female subjects who meet all of the following criteria are eligible for enrollment: 
• 
Are 20 to 49 years of age at the time of enrollment 
• 
Have a documented history of a single yellow fever vaccination with YFVax. 
• 
Not previously vaccinated for tick-borne encephalitis virus (TBEV), Japanese 
encephalitis virus (JEV), or dengue virus. 
• 
Subjects of childbearing potential who are willing to use an acceptable form of 
contraception 30 days prior to the YFV-17D vaccination, and until their participation in this study 
is completed, and who have a negative pregnancy test and express the intent not to conceive a 
child while participating in the study. If the female subject is sexually active and is of 
childbearing potential, she must agree to use an acceptable method of birth control (birth control 
pills, Depo-Provera, Norplant, intrauterine device [IUD], NuvaRing, or diaphragm/condom plus 
spermicide) or remain abstinent. 
 
Exclusion Criteria: 
Subjects who meet any of these criteria are not eligible for enrollment: 
• 
A known history of infection or serologic evidence consistent with prior infection with 
YFV, dengue fever, JEV, or West Nile Virus (WNV). 
• 
Have a history of acute hypersensitivity reaction to any components of the yellow fever 
vaccine (including egg or gelatin). 
 

 
5 
 
 
Title of the Protocol: A phase I/II study of the immunogenicity of the yellow fever vaccine 
17D (YFVax®) in adults with prior 17D vaccination. 
 
• 
any history of progressive or severe neurologic disorder, seizure disorder or neuro-
inflammatory disease (eg, Guillain-Barré syndrome). 
• 
known or suspected impairment/alteration of immune function. 
• 
Known or suspected liver dysfunction. 
• 
serious chronic or progressive disease according to the judgment of the investigator (eg, 
neoplasm, hematologic malignancies, diabetes; cardiac, renal, or hepatic disease). 
• 
a body mass index (BMI) greater than or equal to 35 kg/m2 (=weight in kg/ height in 
meters2). 
• 
women who are pregnant or breastfeeding. 
• 
Subjects of childbearing potential, sexually active with men, and have not used any of 
the acceptable contraceptive methods for at least 2 months prior to vaccination. Subjects who 
fail this screen may initiate or continue acceptable contraception until 2 months of use and then 
rescreen. 
 
Clinical and Laboratory Evaluations: Study subjects will undergo a physical exam prior to 
vaccination and on post-vaccination days 6 and 14. Complete blood count and complete 
metabolic panel tests will be drawn and reviewed prior to vaccination. Vaccination will not 
proceed if CBC, creatine, or liver function tests fall outside the range of normal. Liver function 
tests will be repeated on day 6, day 14, and day 28. Serum will be tested on days 1, 2, 4, 6, 8, 
10, 12, and 14 for vaccine viremia using plaque assay, virus isolation by tissue culture and virus 
genomic copy by quantitative reverse transcription polymerase chain reaction (qRT-PCR). 
Serum will be tested for innate immune response markers will be assayed on days 1, 2, 8, 14 
and 28.  
 
Statistical considerations and power analyses: To execute this aim, we will work with Dr. 
Naleway at KPNW to identify previously vaccinated subjects 8 or more years post vaccination. 
KNPNW vaccinates ~400 individuals/year aged 20-49 years old. Assuming we contact 400 
subjects/year with ~4% uptake, we expect to recruit seventeen subjects/year for 2 years for a 
total of 34 subjects over the study period. Subjects will be recruited pre-vaccination and followed 
for 30 days post-vaccination with blood draws identical to those through Day 30 in Aim 1. We 
assume little to no loss-to-follow up as this study will be conducted over only 60 days, at the end 
of study Year 5 we expect we will have enrolled at least 32 subjects over 2 years to fully 
evaluate.  
Power calculations. Based on prior studies, we assume a priori a sterilizing threshold of NT90 
>1:40. Assuming that 10% of 16 subjects with titers >1:40 boost, and 75% of 16 subjects with 
titers <1:40 boost, against the null that 50% will boost each in group, the study has power=0.85 
(χ2=12.28, DF=3, α=0.05, 32 subjects) to detect a difference in boost between two groups 
divided at an NT90 threshold of >1:40. The study retains power >0.80 with as few as 22 
subjects under the same assumptions, and 20 subjects will have power=0.76. There are no 
prior data available to estimate the proportions of subjects who will develop a detectable viral 
load on 17D re-vaccination, and we consider evaluation for viral load exploratory but extremely 
important for the field. We assume that viral load is most likely to be detected in re-vaccinated 
subjects with NT90<1:40, or 16 (50%) subjects, and we contend that the null hypothesis is that 
no more than 1 in 100 will have a detectable viral load as this is consistent with reasoning 
behind CDC and WHO recommendations for once-in-a-lifetime vaccination. Under these 

 
6 
assumptions, the study has power=0.80 (test of 1 proportion1) to detect a viral load in at least 1 
in 10 subjects. 
 
Human Subjects: 
Vaccine Risks. YFV-17D is Food and Drug Administration (FDA)-approved in the US for both 
primary vaccination in persons aged >6 months traveling to YFV endemic regions and booster 
vaccination in subjects considered high risk for YFV exposure.  YFV-17D is widely   
Title of the Protocol: A phase I/II study of the immunogenicity of the yellow fever vaccine 
17D (YFVax®) in adults with prior 17D vaccination. 
 
17D is acknowledged as one of the safest live attenuated vaccines in use. Over 500 million 
people have been immunized with 17D. YFV-17D has a strong record of safety and tolerability 
dating back to its introduction in 1936. Common reactions to 17D vaccination include mild 
headaches, myalgia, low-grade fevers, or other minor symptoms for 5-10 days. Other symptoms 
may include localized injection site pain, swelling, erythema, or warmth for up to a week after 
vaccination. In one large phase III trial,2 the most common systemic side effects were headache 
(33% of subjects), myalgia (25%), malaise (19%), fever (15%), and chills (11%). The most 
serious and potentially life-threatening AEs related to this vaccine are neurotropic (YEL-AND) 
and viscerotropic disease (YEL-AVD). In the US civilian sector analyzed from 2000 to 2006, 
there were 1.5 million doses of YFV-17D administered. YFV-17D-associated neurotropic 
disease (YEL-AND) has an approximate incidence of 0.8 cases per 100,000 vaccinations (all 
ages) but is not typically lethal. YFV-17D-associated viscerotropic disease (YEL-AVD) has an 
incidence of 0.4 cases per 100,000 vaccinations (all ages) with an approximate 50% mortality 
rate, resulting in up to 2 fatalities per million vaccinations.  There is a reduced incidence of 
approximately 0.07 to 0.24 cases per 100,000 vaccinations in subjects aged 19 to 49.  
 
Risk in booster vaccination: The CDC reports data from observational studies that include 333 
million vaccine doses, with 1,255 SAEs overall; vaccination type (primary vs boost) was 
reported for 201 SAEs, with 14 (7%) of reported SAEs occurring on booster vaccination. CDC 
reports YEL-AND observational data for 462 million vaccine doses distributed with 218 YEL-
AND cases: vaccination type was known for 110 of the YEL-AND cases, of which 3 (3%) were 
booster vaccinations. CDC also reports YEL-AVD observational data for 437 million vaccine 
doses distributed with 72 YEL-AVD cases; vaccination type was known for 31 of the YEL-AVD 
cases, of which 1 (3%) was a booster vaccination. This study has safety lab provisions for early 
detection and management of YEL-AVD. 
 
Blood Draw Risks: Risks associated with drawing blood from the arm include some pain when 
the needle is inserted and a small risk of bruising and/or infection at the place where the needle 
enters the arm. Some people may experience lightheadedness, nausea, and/or fainting. 
 
Benefits. The proposed studies are not expected to be of direct benefit to study subjects. 
However, their participation in these studies may further increase our understanding of the 
mechanisms underlying long term immunity following 17D vaccination. 
  
 
 

 
7 
2 Summary of study events 
 
Visit 
Screening 
visit 
Baseli
ne V1 
V2 
V3 
V4 
V5 
V6 
V7 
V8 
V9 
Visit window (days post 
vaccination) 
up to 30 
days pre-
vaccination 
0 
2  
(±1) 
 
4  
 (±1) 
 
6  
(±1) 
 
8  
(±1) 
 
10 (±1) 
 
12  
(±1) 
 
14  
(±1) 
 
28  
(±7) 
 
Informed consent  
X 
 
 
 
 
 
 
 
 
 
Assessment of eligibility 
criteria  
X 
 
 
 
 
 
 
 
 
 
Demographics, medical 
history, and prior 
medication/ vaccination 
X 
 
 
 
 
 
 
 
 
 
Concomitant 
medications/vaccinations 
during study period 
X 
 
 
 
 
 
 
 
 
 
Travel History 
Questionnaire 
X 
 
 
 
 
 
 
 
 
 
Urine Pregnancy Test (if 
applicable) 
X 
 
 
 
 
 
 
 
 
 
Blood sample for plasma 
and peripheral blood 
mononuclear cells 
 
20 mL 
 
 
10 mL 
 
 
 
10 mL 
10 mL 
Blood sample for serum 
viremia and/or innate 
immune response as 
specified in the protocol2  
 
3 mL 
3 mL 
3 mL 
3 mL 
3 mL 
3 mL 
3 mL 
3 mL 
 
Blood sample for serology 
(screening and post-
vaccination antibodies) 
5.0 mL 
 
 
 
 
 
 
 
 
5.0 mL 
Blood sample for CBC 
3.0 mL 
 
 
 
 
 
 
 
 
 
Blood sample for CMP 
4.0 mL 
 
 
 
 
 
 
 
 
 
Blood sample for 
Paxgene4 
 
2.5 mL 
 
 
 
 
 
 
 
 
Total Blood Volume 
12.0 mL 
25.5 
mL 
3 mL 
3 mL 
13 mL 
3 mL 
3 mL 
3 mL 
13 mL 
15 mL 
Study Diary 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
YFV Vaccination 
 
X 
 
 
 
 
 
 
 
 
Full Investigator 
examination 
X 
X 
 
 
 
 
 
 
 
X 
Focused investigator 
examination as needed 
 
 
X 
X 
X 
X 
X 
X 
X 
 
 
 
 
 

 
8 
3 
TABLE OF CONTENTS 
  
PROTOCOL TITLE PAGE 
 
 
 
 
 
 
 
 
  2 
1 
SYNOPSIS 
 
 
 
 
 
 
 
 
 
  3 
2 
SUMMARY OF STUDY EVENTS 
 
 
 
 
 
 
  6 
3 
TABLE OF CONTENTS 
 
 
 
 
 
 
 
  7 
4 
ABBREVIATIONS 
 
 
 
 
 
 
 
 
10 
5 
BACKGROUND AND RATIONALE  
 
 
 
 
 
12 
5.1 
INTRODUCTION 
 
 
 
 
 
 
 
 
12 
5.2 
STUDY RATIONALE  
 
 
 
 
 
 
 
12 
5.3 
RATIONALE FOR SELECTION OF STUDY POPULATION  
 
 
15 
5.4 
SAFETY AND RISKS OF YFV-17D IN HUMANS 
 
 
 
 
15 
5.5 
POTENTIAL BENEFITS 
 
 
 
 
 
 
 
16 
5.6 
SUMMARY OF CLINICAL SAFETY DATA  
 
 
 
 
16 
6 
STUDY OBJECTIVES AND PURPOSE 
 
 
 
 
 
16 
6.1 
PRIMARY OBJECTIVE 
 
 
 
 
 
 
 
17 
6.2 
SECONDARY OBJECTIVE  
 
 
 
 
 
 
17 
7 
STUDY DESIGN 
 
 
 
 
 
 
 
 
17 
7.1 
DESCRIPTION OF STUDY DESIGN 
 
 
 
 
 
17 
7.2 
STUDY ENDPOINTS  
 
 
 
 
 
 
 
17 
7.2.1 Primary Endpoint. 
 
 
 
 
 
 
 
 
17 
7.2.2 Secondary Endpoints  
 
 
 
 
 
 
 
17 
7.2.3 Exploratory Endpoints 
 
 
 
 
 
 
 
17 
7.2.4 Safety Endpoints 
 
 
 
 
 
 
 
 
18 
7.3 
CRITERIA FOR PREMATURE TERMINATION OF THE STUDY  
 
18 
8 
SELECTION AND WITHDRAWAL OF SUBJECTS  
 
 
 
18 
8.1 
INCLUSION CRITERIA 
 
 
 
 
 
 
 
18 
8.2 
EXCLUSION CRITERIA 
 
 
 
 
 
 
 
18 
8.3 
CRITERIA FOR DELAY 
 
 
 
 
 
 
 
20 
8.4 
EARLY TERMINATION OF SUBJECT TRIAL PARTICIPATION 
 
 
20 
9 
TREATMENT OF SUBJECTS 
 
 
 
 
 
 
20 
9.1 
DESCRIPTION OF STUDY VACCINE AND DOSING REGIMEN  
 
20 
9.1.1 Interruption of Study Drug 
 
 
 
 
 
 
 
21 
9.1.2 Vaccine Toxicity Management 
 
 
 
 
 
 
21 
9.2 
DOCUMENTATION OF STUDY PRODUCT  
 
 
 
 
21 
9.3 
SUBJECT COMPLIANCE 
 
 
 
 
 
 
 
22 
10 
STUDY PROCEDURES 
 
 
 
 
 
 
 
22 
10.1 
TRIAL PROCEDURES 
 
 
 
 
 
 
 
22 
10.1.1 Informed Consent 
 
 
 
 
 
 
 
 
22 
10.1.2 Demographics, Medical History and prior medications 
 
 
 
22 
10.1.3 Documentation of Trial Entrance/Randomization 
 
 
 
 
23 
10.1.4 Physical Examination  
 
 
 
 
 
 
 
23 
10.1.5 Vital Signs 
 
 
 
 
 
 
 
 
 
23 
10.1.6 Immunogenicity Assessment  
 
 
 
 
 
 
23 
10.1.7 Processing, Labeling and Storage of Biological Samples 
 
 
 
23 
10.1.8 Safety Assessments  
 
 
 
 
 
 
 
23 
10.1.9 Clinical Safety Laboratory Variables  
 
 
 
 
 
23 
10.1.10 
Contraception and pregnancy Avoidance Procedures 
 
 
24 
10.2 
MONITORING SUBJECT COMPLIANCE 
 
 
 
 
 
24 
10.3 
SCHEDULE OF OBSERVATIONS AND PROCEDURES 
 
 
 
24 
10.3.2 Pre-vaccination procedures  
 
 
 
 
 
 
24 

 
9 
10.3.3 Vaccination procedures 
 
 
 
 
 
 
 
24 
10.3.4 Post vaccination procedures  
 
 
 
 
 
 
24 
10.3.5 Site visits after CAccination: Days 2, 4, 6, 8, 10, 12, 14, 28  
 
 
25 
10.3.6  Final Visit 
 
 
 
 
 
 
 
 
 
26 
10.4 
SAMPLE COLLECTION 
 
 
 
 
 
 
 
26 
10.4.1 Blood Samples 
 
 
 
 
 
 
 
 
26 
10.4.2 PBMC Cryopreservation 
 
 
 
 
 
 
 
26 
11 
SAFETY ASSESSMENTS 
 
 
 
 
 
 
 
26 
11.1 
SAFETY ASSESSMENTS OVERVIEW 
 
 
 
 
 
26 
11.2 
DEFINITIONS  
 
 
 
 
 
 
 
 
27 
11.2.1 Adverse Events 
 
 
 
 
 
 
 
 
27 
11.2.2 Medically Attended Adverse Events  
 
 
 
 
 
28 
11.2.3 Serious Adverse Events 
 
 
 
 
 
 
 
28 
11.2.4 Unexpected Adverse Events  
 
 
 
 
 
 
28 
11.3 
COLLECTION AND RECORDING OF ADVERSE EVENTS 
 
 
29 
11.3.1 Collection Period 
 
 
 
 
 
 
 
 
29 
11.3.2 Collection of Adverse Events  
 
 
 
 
 
 
29 
 11.3.3 Recording Adverse Events 
 
 
 
 
 
 
 
29 
11.3.4 Recording Serious Adverse Events  
 
 
 
 
 
30 
11.4 
GRADING AND ATTRIBUTION OF ADVERSE EVENTS 
 
 
 
30 
11.4.1 Grading Criteria 
 
 
 
 
 
 
 
 
30 
11.4.2 Attribution Definitions  
 
 
 
 
 
 
 
30 
11.5 
REPORTING OF ADVERSE EVENTS 
 
 
 
 
 
31 
11.5.1 Adverse Events Requiring Expedited Reporting 
 
 
 
 
31 
11.5.2 Reporting Timeline. 
 
 
 
 
 
 
 
 
31 
11.5.3 Reporting of Adverse Events  
 
 
 
31 
11.5.4 Reporting Pregnancy as a Serious Adverse Event  
 
 
 
31 
11.6 
REVIEW OF SAFETY INFORMATION 
 
 
 
 
 
32 
11.6.1 Routine ISM Review  
 
 
 
 
 
 
 
32 
11.6.2 ISM Product Safety Associate Review 
 
 
 
 
 
32 
11.6.3 Independent Safety Monitor  
 
 
 
 
 
 
32 
11.6.4 Institutional Review Board Reporting of Adverse Events 
 
 
 
32 
11.7 
PROTOCOL DEVIATIONS 
 
 
 
 
 
 
 
32 
11.8 
CLINICAL MONITORING 
 
 
 
 
 
 
 
33 
12 
DATA HANDLING AND RECORD KEEPING 
 
 
 
 
33 
12.1 
EDC AND SOURCE DOCUMENTATION 
 
 
 
 
 
33 
12.2 
DATA MANAGEMENT 
 
 
 
 
 
 
 
33 
12.3 
DATA MONITORING  
 
 
 
 
 
 
 
33 
12.4 
RECORD RETENTION 
 
 
 
 
 
 
 
33 
13 
STATISTICAL ANALYSIS 
 
 
 
 
 
 
 
34 
 
13.1 
ANALYSIS POPULATIONS  
 
 
 
 
 
 
34 
13.1.1 Analysis Population 
 
 
 
 
 
 
 
 
34 
13.2 
STATISTICAL METHODS 
 
 
 
 
 
 
 
34 
13.2.1 Demographics, Baseline Characteristics, and Disposition  
 
 
34 
13.2.2 Primary Analysis – 
 
 
 
 
 
 
 
 
34 
13.2.3 Secondary Analysis  
 
 
 
 
 
 
 
35 
13.2.4 SAFETY DATA 
 
 
 
 
 
 
 
 
35 
13.2.5 SAFETY REVIEW 
 
 
 
 
 
 
 
 
35 
13.3 
SAMPLE SIZE 
 
 
 
 
 
 
 
 
35 
14 
HUMAN SUBJECTS PROTECTION  
 
 
 
 
 
36 
14.1 
DISCONTINUATION OF STUDY 
 
 
 
 
 
 
36 

 
10 
14.2 
ETHICS 
 
 
 
 
 
 
 
 
 
36 
14.2.1 Good Clinical Practice and Institutional Review Board Review 
 
 
36 
14.2.2 Informed Consent 
 
 
 
 
 
 
 
 
37 
14.3 
STUDY RISKS 
 
 
 
 
 
 
 
 
37 
14.3.1 Vaccine Risks  
 
 
 
 
 
 
 
 
37 
14.3.2 Blood Draw Risks 
 
 
 
 
 
 
 
 
37 
14.4 
STUDY BENEFITS 
 
 
 
 
 
 
 
 
37 
14.5 
CONFIDENTIALITY  
 
 
 
 
 
 
 
37 
14.6 
DISCLOSURE OF DATA 
 
 
 
 
 
 
 
38 
14.7 
PUBLICATION OF RESEARCH FINDINGS  
 
 
 
 
38 
15 
SUBJECT COMPENSATION  
 
 
 
 
 
 
38 
16 
REFERENCES 
 
 
 
 
 
 
 
 
38 
 
 

 
11 
4 ABBREVIATIONS 
 
ACIP Advisory Committee on Immunization Practices 
AE 
Adverse Event 
AUC 
Area Under the Curve 
BMI 
Body Mass Index 
CDC 
Centers for Disease Control and Prevention 
CFR 
Code of Federal Regulations 
CI 
Confidence Interval 
CPT 
Cell Preparation Tube 
CO2 
Carbon Dioxide 
CTL 
Cytotoxic T-Lymphocyte 
EDC 
Electronic Data Capture 
ELISA Enzyme-Linked Immunosorbent Assay 
FACS Fluorescence-Activated Cell Sorting 
FDA 
Food and Drug Administration 
GCP 
Good Clinical Practice 
GMP Good Manufacturing Practice 
HEP 
Hepatitis 
HIV 
Human Immunodeficiency Virus 
ICH 
International Conference on Harmonization 
ID 
Identification 
Ig 
Immunoglobulin 
IND 
Investigational New Drug 
IRB 
Institutional Review Board 
ISM  
Independent Safety Monitor 
IUD 
Intrauterine Device 
IVIG 
Intravenous Immunoglobulin 
JEV 
Japanese Encephalitis Virus 
LNI 
Log10 Neutralization Index 
MedDRA 
Medical Dictionary for Regulatory Activities 
NIH 
National Institutes of Health 
NIAID National Institute of Allergy and Infectious Diseases 
OHSU Oregon Health and Science University 
PRNT Plaque Reduction Neutralization Test 
PBMC Peripheral Blood Mononuclear Cell 
PFU 
Plaque Forming Unit(s) 
PI 
Principal Investigator 
RNA 
Ribonucleic Acid 
RT-PCR 
Reverse Transcriptase-Polymerase Chain Reaction 
SAE 
Serious Adverse Event 
SC 
Subcutaneous(ly) 
SD 
Standard Deviation 
SDCC Statistical and Data Coordinating Center 
SSL 
Secure Sockets Layer 
TBEV Tick-Borne Encephalitis Virus 
US 
United States (of America) 
VAERS 
Vaccine Adverse Event Reporting System 
WNV West Nile Virus 
YEL-AND 
Yellow Fever Vaccine-Associated Neurotropic Disease Y 
EL-AVD 
Yellow Fever Vaccine-Associated Viscerotropic Disease  

 
12 
YF 
Yellow Fever 
YFV 
Yellow Fever Virus 
YFV-17D 
Yellow Fever Vaccine Virus from Sanofi Pasteur (YF-VAX®) 
YF-VAX® 
Yellow Fever Vaccine Manufactured by Sanofi Pasteur 
 
 
 

 
13 
5 
BACKGROUND AND RATIONALE 
 
5.1 
Introduction 
 
Yellow fever virus (YFV) is the prototype flavivirus, closely related to dengue (DENV), Zika 
(ZIKV) and Japanese encephalitis (JEV) viruses. YFV is transmitted primarily by Aedes aegypti 
mosquitoes in urban transmission cycles and by Haemogogus spp. forest canopy mosquitoes in 
sylvatic, non-human primate transmission cycles. Historically YFV has been the most important 
arthropod-borne viral pathogen of humans worldwide, causing explosive epidemics in Europe, 
the Americas and Africa in the 18th, 19th and early 20th centuries,1,2 and it remains endemic to at 
least 34 African and 13 South American countries. Control of YFV has been accomplished 
largely through vector abatement and mass vaccination with the YFV vaccine 17D.2 In the early 
2000’s the WHO estimated that there were approximately 200,000 cases/year, with 30,000 
deaths,3 but in the past decade falling vaccination rates and spillover of sylvatic YFV have led to 
a dramatic resurgence of YFV disease, with an estimated 130,000 severe cases and 78,000 
fatalities in Africa in 2013,4 an estimated 7,334 sylvatic-urban cases in Angola and the 
Democratic Republic of Congo in 2015-2016,6 and a series of sylvatic outbreaks in Brazil since 
2016 that threaten to spill into urban settings.7,8 Ominously, urban city parks in Sao Paolo were 
closed in 2017 after at least one park-dwelling monkey died from YFV9, while the Republic of 
Congo reported at least once confirmed case of YFV in the heart of the port city of Porte Noire 
in July, 201810. Additionally, 11 YFV cases were imported by travelers into China from Angola in 
2016,11 potentially exposing the largely unvaccinated Asian population to epidemic 
transmission. Taken together, the rising incidence of YFV outbreaks in previously well-controlled 
endemic settings and introductions into unvaccinated populations in endemic and non-endemic8 
settings corroborate the enormous risk YFV still poses to the global population. 
Since its development by Max Theiler in 1937, the live-attenuated virus (LAV) YFV vaccine 
17D—administered via population-wide vaccination—has been the cornerstone of YFV control. 
YFV-17D is the parental vaccine strain from which all current YFV vaccines are derived; there 
are three sub-strains used by global manufacturers, 17D-204 (YF-Vax® in US, Stamaril® in 
EU), 17D-213 in and 17DD in Latin America,12 which have been found to be equivalent in  
trials.12,13 Despite being widely regarded as highly immunogenic and durable, historically the 
vaccine has been administered as a prime followed by boosting every 10 years. However, the 
rise in global demand for vaccine doses because of recent YFV outbreaks has resulted in 
unprecedented worldwide shortages of 17D.14 These critical shortages have led to strategies to 
extend vaccine stocks, including a directive by the CDC and WHO to replace the long-standing 
10-year boost with once-in-a-lifetime vaccination for most vaccinees, and global and national 
health organizations (WHO, Brazilian MOH) exploring fractional rather than full-dose 
vaccination.1 The policy changes should relieve short-term pressure on the vaccine supply, but 
they move 17D vaccination policies into uncharted waters. Scrutiny and criticism of these 
strategies, particularly that of dropping the 10-year boost, make clear that definitive studies to 
validate or revise these strategies have not been done. Specifically, the immune correlates of 
protection, how the vaccine induces protective immunity, and who maintains and who loses 
long-term protection have not been well-defined. 
 
5.2 
Rationale  
 
In 1973 Mason et al14 demonstrated a clear correlation between neutralizing antibody titers and 
protection against lethal challenge with the virulent YFV Asibi strain at 20 weeks post-
vaccination with 17D in Rhesus macaques. Subsequently, neutralizing antibody titers have 
become the most widely accepted correlate of 17D-induced protection and are believed to be 
necessary and sufficient for YFV immunity.15 Generalizability of Mason’s data is difficult, 

 
14 
however, because Mason’s protective threshold titer was based on a ratio between log10 pre- 
and post-vaccination neutralizing antibody titers – the log neutralizing index (LNI) –equivalent 
LNI or alternate protective neutralizing antibody titer has never been validated in humans for 
symptomatic or lethal wild-type infection. The sporadic, deadly nature of YFV outbreaks, the 
remote settings in which they often occur, and the logistic challenges of obtaining pre-
vaccination titers in vaccinees in endemic areas effectively preclude prospective human studies 
that could determine an antibody titer that protects against wild-type YFV disease. 
Nevertheless, 90% percent plaque reduction neutralization test (PRNT90 or NT90) titers >1:10 
are often taken as an immune correlate of protection16, despite a lack of supporting data.  
 
A limited number of studies have used 17D booster doses as a live-attenuated challenge model 
(boosting is a true homologous live-virus challenge) to evaluate neutralized antibody titers that 
confer differing degrees of immunity. In these studies, “sterilizing” immunity is defined as a less 
than four-fold rise in neutralizing antibody titer post-boost and/or no detection of vaccine virus by 
virus isolation or PCR.  These studies suggest that NT80 or NT90 titers >1:40 are probably 
required for reliable sterilizing immunity against live attenuated 17D challenge (Table 1). In 1998 
Reinhardt et al.17 boosted 5 subjects >10 years post first vaccination. All 5 subjects had pre-
boost NT90 titers ranging from ~1:45 to 1:90 and none had a four-fold or greater boost in NT90  
post-boost and none had a detectable viral load by PCR. While a threshold-sterilizing NT was 
not identified, as all five subjects were fully protected, Reinhardt showed that 17D elicits durable 
sterilizing immunity correlated with an NT90 at of least >1:45. In a much larger study, Hepburn et 
al.18 surveyed 1029 laboratory technicians working with wild-type YFV from 1976-2002 using 
NT80 against the wild-type YFV Asibi strain. When a surveillance NT80 dropped <1:40 or a fixed 
number of years had passed, the lab worker was boosted and had follow-up serology.  Among 
subjects with NT80 >1:40, 71/79 (90%) had sterilizing immunity. This proportion dropped to 
42/121 (35%) for subjects with NT80 between 1:10 and 1:40, and only 183/829 (22%) of subjects 
with NT80 <1:10 (Table 5.1) had sterilizing immunity. Hepburn et al. had two critical findings: 1) 
there was a clear “dose-response” relationship between pre-boost NT80 and loss of sterile 
immunity, and 2) substantial evidence that YFV immunity wanes over time, with 829/1029 
subjects NT80 titers vs Asibi falling to <1:10 over the time period studied. Most recently, 
Kongsgaard et al.19 conducted a prospective study of 28 Danish individuals presenting for 17D 
boost 9-36 years after primary vaccination. Pre-boost, 23 of the 28 subjects had 50% 
neutralization titers (NT50) >1:10 (NT90 titers were not reported), while the remaining five had 
NT50 <1:10. Overall, 21/28 subjects boosted, with 78% (18/23) in the >1:10 group and 3/5 in the 
<1:10 group. While Kongsgaard et al.19 did not further stratify by NT50 or report individual 
results, they reported that higher pre-boost NT50 titers significantly correlated with less-than-
four-fold rises in NT50 titers 18. Importantly, this study demonstrated that less stringent NT50 titers 
>1:10 are neither above a threshold of protection nor predictive of sterilizing immunity. 
 
Study 
Threshold 
Sterile 
immunity/total N 
% Sterile immunity 
Reinhardt et al., 
PRNT90 >1:40 
5/5 
100% 
Hepburn et al., 
PRNT80 >1:40 
71/79 
90% 
Hepburn et al., 2002 
PRNT80 >1:10 
42/121 
35% 
Hepburn et al., 2002 
PRNT80 <1:10 
183/829 
22% 
Kongsgaard et al., 
PRNT50 >1:10 
5/23 
22% 
Kongsgaard et al. 
PRNT50<1:10 
2/5 
40% 
Table 5.1. Summary of studies describing proportions of subjects with sterilizing immunity (<4-
fold rise between pre- and post-boost in antibody titer and/or no detection of vaccine virus by irus 
isolation or PCR) on 17D re-vaccination. 

 
15 
The clinical significance of loss of YFV-neutralizing antibodies has become increasingly 
apparent: while the CDC 2015 recommendations reported only 18 confirmed and documented 
cases of 17D vaccine failure38 out of >540 million administered 17D doses, more recent 
observational data suggest that vaccine failure occurs much more frequently. The Pan 
American Health Organization (PAHO) has made publicly available confirmed YFV disease 
cases for Peru, Brazil, Columbia and Bolivia from 2000-2014.39 Of 1071 cases, 61% (653/1071) 
had not been vaccinated, 8% (85/1071) had been vaccinated, and an additional 31% (333/1071) 
had unknown vaccine status. This last proportion is expected to include additional vaccine 
breakthrough cases.  A separate study of sylvatic YFV in Brazil25 evaluated 831 YFV cases from 
1972-2008 reported to the Ministry of Health, finding that 45% (372/831) of reported cases had 
no recorded vaccine status but the remaining 55% (459/831) had a recorded status, of whom 
only three had been vaccinated <10 years earlier, but the remaining 456 reportedly were 
vaccinated >10 years before infection.2,25 While limited, these data strongly suggest loss of YFV 
neutralizing antibodies is clinically important, even lethal, and that the risk increases for some 
proportion of vaccinees 10 our more years post-vaccination. 
 
Before changing 17D vaccination recommendations from 10-year boost to once-in-a-lifetime 
vaccination, the CDC reviewed prior research on long- term YFV immunity comprised of 13 
cross-sectional serosurveys, finding an average of 88% of vaccinees remain seropositive for 
YFV-neutralizing antibodies (Table 3), which suggests the 10-year boost was not necessary. 
However, careful review of this prior research shows substantial heterogeneity in both methods 
and results, calling into question the overall conclusion, including: 1) the conflation of human 
antibody titers and mouse protection studies, 2) the conflation of immunity in endemic and non-
endemic populations, and 3) treating NT titers with different neutralization cutoffs as 
comparable2. Specifically: four29-32 of the 13 studies were mouse protection studies for which no 
correlation of protection in humans has been reasonably established2; of the remaining human 
studies, four were conducted in endemic countries33-36 and five in non-endemic countries 
17,26,27,37,38. When stratified and pooled by endemic/non-endemic studies, 92.7% (359/387) of 
endemic subjects were seropositive, but only 83.7% (282/337) of non-endemic subjects were 
seropositive. This disparity is highly statistically significant (P<0.001, χ2 test). This disparity may 
be due to unreported YFV vaccine boosts in endemic subjects, infection with other flaviviruses 
in endemic subjects, age differences in vaccinees, or other unknown confounders/effect 
modifiers. Irrespective, these data argue that endemic and non-endemic populations should not 
be conflated, with non-endemic studies representing the worst-case scenario for vaccine 
efficacy. Once segregated by endemic/non-endemic origin and adjusted for differences in 
neutralizing antibody titer thresholds, we estimate that at least one in five, ~20%, vaccinees 
become seronegative by 10 or more years post-vaccination, while the remainder appear to 
maintain life-long protective neutralizing antibody titers (Figure 1, Wanjeri et al. accepted, 
Journal of Infectious Diseases). The finding that one in five 17D vaccinees may lose protective 
neutralizing antibodies by 10 or more years post-vaccination has significant implications for both 
vaccine deployment and vaccinee safety, but also offers a unique opportunity to compare and 
contrast 17D immunity over time between seronegative and seropositive subjects and define the 
determinants of long-term 17D serostatus.  
 
Taken together, we find: 1) substantial evidence that neutralizing antibodies play the critical role 
in protection against YFV infection following vaccination, 2) at least one in five 17D vaccinees 
living in non-endemic countries will lose protective neutralizing antibody titers, for which there 
are, 3) potentially substantial safety and public health implications. However, the studies 
supporting these preliminary conclusions are limited by their retrospective and cross-sectional 
design, providing no knowledge of why or when vaccinees become seronegative, or the clinical 
and epidemiologic implications of seronegative status.  

 
16 
 
The rationale for this trial is largely established by the Hepburn et al.18 study that provided 
substantial data supporting the use of 17D boost as a challenge model for sterilizing immunity. 
However, Hepburn has several key shortcomings that limit generalizability to other vaccinee 
populations: 1) it was not specifically designed to test immunity at 10 years post prime, but 
rather assessed boost when neutralizing antibody titers dropped below a certain level, 2) many 
subjects had received more than 1 boost, 3) non-standard wild-type Asibi strain was used in the 
PRNT assay and an 80% rather than 90% neutralization cut-off was reported, and 4) vaccine 
viremia was not assessed. While measuring boost in neutralization titer has been a standard 
approach to characterizing strength of protective immunity from 17D vaccination, measuring 
viral load on boost has been an overlooked but critically important metric for characterizing long-
term immunity. The Hepburn study was retrospective and not positioned to assess vaccine viral 
load in boosted lab workers, but the Kongsgaard study,18  although done in a prospective 
manner, did not assess viral load in vaccinees in the acute 1- to 14-day post-vaccination period. 
Reinhardt et al. did assess viral load, and found none, but the study was quite small and only 
looked at vaccinees with neutralization titers >1:40, a titer that, based on Hepburn et al., 
appears to confer sterilizing immunity. A detectable viral load in a boosted vaccinee suggests 
both the potential for transmissible or symptomatic breakthrough viremia and is the only way to 
assess whether “undetected” T-cell or low-level neutralizing antibodies have the potential to 
protect in “seronegative” vaccinees or if these vaccinees with undetectable titers are essentially 
“naïve.” 
 
Drawing upon the model that 17D boost is a homologous live-virus challenge, we will 
prospectively recruit and boost 35 individuals with history of 1 prior 17D vaccination ~10 years 
prior. We will characterize pre- and post-boost neutralizing antibody titers, boost vaccine viral 
load, and innate and B-cell responses in boosted vaccinees.  We will identify titers below which 
vaccine viral load and/or antibody boost responses are detected and higher titers that confer 
sterilizing immunity without detectable viral load or antibody titer boost, establishing both 
stringent and relaxed correlates of antibody protection. We will also assess whether some 
vaccinees with undetectable neutralizing antibodies are still protected against 17D challenge, 
setting the stage to explore the extent to which protection may be provided by other immune 
mechanisms. 
 
5.3 
Rationale for Selection of Study Population 
 
To minimize study risks, this study will not include any vulnerable populations (i.e. prisoners, 
pregnant women, children, neonates, and/or adults lacking capacity). This study will involve 
healthy subjects 20 to 49 years of age with documented history of prior yellow fever vaccination. 
This age range for inclusion was determined based on low susceptibility of young to middle 
aged adults to SAEs following YFV-17D vaccination (See 5.5) below.  YFV-17D is an FDA-
approved vaccine but it is contraindicated for vaccination of infants younger than 9 months of 
age because of an increased risk of encephalitis. It is also contraindicated in 
immunosuppressed subjects and subjects with known egg hypersensitivity (see YF-VAX® 
package insert for details).  YFV-17D vaccination is generally considered safe for adults, with 
the most common side effects being flu-like symptoms. However, rare cases of serious or lethal 
infections stemming from YFV-17D vaccination have been reported. More than 500 million 
people have been immunized with this vaccine worldwide, with an estimated mortality rate of 
0.04 to 2.1 deaths/million doses administered. However, these statistics include individuals of all 
ages (including the young and elderly who are more susceptible to SAEs) and combine primary 
and booster vaccinations. 
 

 
17 
5.4 
Safety and Risks of YFV-17D In Humans 
 
17D has a strong record of safety and tolerability dating back to its introduction in 1936. Safety 
data suggest that booster vaccination as at least 10-fold lower risk of a vaccine related severe 
adverse event (SAE) compared to primary vaccination. The CDC reports data74 from 
observational studies that include 333 million vaccine doses, with 1,255 SAEs overall; 
vaccination type (primary vs boost) was reported for 201 SAEs, with 14 (7%) of reported SAEs 
occurring on booster vaccination. The most serious and potentially life-threatening AE related to 
this vaccine are neurotropic (YEL-AND) and viscerotropic disease (YEL-AVD). CDC reports 
YEL-AND observational data74 for 462 million vaccine doses distributed with 218 YEL-AND 
cases: vaccination type was known for 110 of the YEL-AND cases, of which 3 (3%) were 
booster vaccinations. CDC also reports YEL-AVD observational data for 437 million vaccine 
doses distributed with 72 YEL-AVD cases;74 vaccination type was known for 31 of the YEL-AVD 
cases, of which 1 (3%) was a booster vaccination. Subjects will be screened for reported 
adverse reactions at first 17D vaccination and excluded if positive. 
 
5.5 
Potential Benefits 
 
Subjects will not personally benefit from participating in this clinical study, but there may be 
scientific benefit or benefit to the population at large. Although each subject will receive a YFV-
17D vaccination booster, there is no guarantee that the booster will have a protective effect 
against YFV following administration. Subjects will be counseled to follow up with their primary 
care physician or a travel clinic for clinical travel recommendations prior to travel to a YFV 
endemic region. 
 
5.9 
Summary of Clinical Safety Data 
 
1) 
YFV-17D is an FDA-approved vaccine introduced in 1936 and has a long history of 
safety in humans. Each year, it is routinely administered by the SC route to more than 200,000 
civilians and more than 500,000 military personnel in the US and has been administered to 
more than 500 million people worldwide. 
2) 
The most common AEs are mild flu-like symptoms. 
3) 
Rare but severe and/or life-threatening allergic reactions (including anaphylaxis) are 
estimated to occur at a rate of ≤ 17 cases per million vaccinations in primary vaccination and 
10-fold less often in booster vaccination. 
4) 
Rare but severe and potentially life-threatening neurological disease (YFV-AND) 
requiring hospitalization is estimated to occur at a rate of 0.8 cases per 
100,000 doses (all age groups) but is typically not lethal. There is a reduced risk of 
approximately 0.15 cases per 100,000 vaccinations in subjects aged 19 to 49. Booster doses 
account for ~3% of all cases. 
5) 
Rare but severe and life-threatening viscerotropic disease (YEL-AVD) requiring 
hospitalization is estimated to occur in 0.4 cases per 100,000 vaccinations (all age groups) with 
an approximate 50% mortality rate, resulting in up to 2 fatalities per million vaccinations.  There 
is a reduced incidence of approximately 0.07 cases per 100,000 vaccinations (9) to 0.24 cases 
per 100,000 vaccinations in subjects aged 19 to 49.  Booster doses account for ~3% of all 
cases.  
 
6 
STUDY OBJECTIVES AND PURPOSE 
 

 
18 
The objective of this study is to use live attenuated yellow fever vaccine 17D as a live virus 
challenge to assess to mechanisms of immune protection in previously vaccinated adults aged 
20-49 years old.  
 
6.1 
Primary Objective 
 
To determine the pre-boost neutralizing antibody titer that correlates with sterile immunity, the 
most stringent measure of vaccine efficacy. 
 
6.2 
Secondary Objectives 
 
To determine whether there is a measurable pre-vaccination antibody titer range in which 
vaccinees experience a memory antibody response as demonstrated by a four-fold rise in pre 
and post-vaccination antibody titers. 
To determine whether there is an antibody titer below which subjects will develop a detectable 
vaccine viremia. 
 
6.3  
Exploratory Objectives 
 
 To determine which innate and adaptive immune responses are stimulated when a pre-immune 
subject is challenged with attenuated live yellow fever virus. To determine whether the 
stimulated immune responses correlate with antibody titer boosting or with detectable viremia. 
 
7 
STUDY DESIGN 
 
7.1 
Description of Study Design 
 
This is a prospective single arm study of the immunogenicity of the yellow fever vaccine 17D 
(YFVax®, Sanofi Pasteur, Swiftwater, PA) in adults who have documented history of prior 
yellow fever vaccination 8 or more years prior. Study subjects will receive a single YFV-17D 
vaccination and be followed for 30 days after vaccination with blood samples collected for pre-
screening and on vaccination days 0, 2, 4, 6, 8, 10, 12, 14 and 28 post vaccination. The study is 
designed to enroll 34 vaccinees at a single site, OHSU. Study duration is expected to be no 
longer than 60 days.  Subject recruitment will be completed in 2 years.   
 
7.2 
Study Endpoints 
 
7.2.1 Primary Endpoint 
 
The primary endpoint will be fold change YFV neutralizing antibody titers levels measured on 
Day 0 and Day 28 post-vaccination. 
 
7.2.2 Secondary Endpoints 
 
• 
Vaccine viremia as measured by qRT-PCR, focus forming assay a virus isolation on days 0, 
2, 4, 6, 8, 10, 12, and 14. 
7.2.3 Exploratory Endpoints 
• 
Frequency and magnitude CD4+ and CD8+ cell populations on day 0, 2, 8, 14, and 28 post 
vaccination.  

 
19 
• 
Multiplexed cytokine levels for 48 different cytokines measured on days 1, 2, 8, 14, and 28 
days post vaccination. 
• 
Titer of vaccination virus preparation as measured by qRT-PCR, focus forming assay and 
virus isolation. 
• 
YFV specific MBC frequency for subject PBMCs collected on days 1, 2, 8, 14, and 28 days 
post vaccination.  
7.2.4 Safety endpoints 
• 
Frequency and percentage of subjects with solicited injection site reactions for 7 days post 
vaccination and solicited systemic events for 14 days post vaccination.  
• 
Frequency and percentage of subjects with any unsolicited Adverse Events (AEs) up to 28 
days post vaccination. 
• 
Frequency and percentage of subjects with Medically Attended Adverse Events (MAAEs) 
throughout the trial. 
 
7.3 
Criteria for Premature Termination of the Study:  Stopping Rules 
 
Study enrollment and vaccination will be suspended pending review of all pertinent data after 
the occurrence of: 
• 
1 Grade 4 or 2 Grade 3 AEs possibly related to the vaccination 
• 
1 SAE possibly related to the vaccination (e.g. viscerotropic or neurological disease) 
Adverse events will be graded according to the Toxicity Grading Scale for Healthy Adult and 
Adolescent Volunteers Enrolled in Preventive Vaccine Clinical Trials guidelines as outlined on 
the FDA website: http://fda.gov/cber/gdlns/toxvac.htm. 
If the study is stopped due to meeting the above criteria, it may not be resumed until all 
pertinent data is reviewed by the Medical Monitor. 
 
8 
SELECTION AND WITHDRAWAL OF SUBJECTS 
 
All entry criteria, including test results, need to be confirmed prior to first vaccination. 
 
8.1 Inclusion Criteria 
 
Subject eligibility is determined according to the following criteria: 
1. Aged ≥20 to <50 years. 
2. Male or female. 
3. In good health at the time of screening as determined by medical history, physical 
examination, and clinical judgement of the investigator. 
4. Documented history of Yellow fever vaccination 8 or more years prior. Documentation 
must be on a primary (not copied) vaccination card or a fully completed electronic 
medical record entry including date of administration and lot number administered. 
5. Subjects who can comply with all trial procedures and are available for the duration of 
follow-up. 
 
8.2 Exclusion Criteria 
 
Any subject who meets any of the following criteria will not qualify for study enrollment: 

 
20 
1. A clinically active infection or self-reported body temperature ≥38°C (100.4°F) within 3 days 
of scheduled date of vaccination (consider whether the finding is an exclusion criterion or 
criterion for delay of vaccination see Section 8.3). 
2. A known hypersensitivity or allergy to any of the trial vaccine components including eggs. 
3. Behavioral/cognitive impairment that, in the investigator’s opinion, may interfere with the 
subject’s ability to participate safely in the trial. 
4. Any history of neurologic disorder, seizure disorder or neuro-inflammatory disease. 
5. Any illness, or history of any illness that, in the investigator’s opinion, could interfere with the 
trial or pose an additional risk to the subject during the trial period. 
6. Known or suspected impairment/alteration of immune function, including: 
a) Chronic use of oral steroids within 60 days prior to enrollment. Inhaled steroids are 
allowed. 
b) Receipt of parenteral steroids within 60 days prior to screening visit.  
c) Receipt of immunoglobulins and/or any blood products within the 3 months prior to 
enrollment or planned receipt during the trial. 
d) Receipt of immunostimulants within 60 days prior to screening visit 
e) Immunosuppressive therapy such as anti-cancer chemotherapy or radiation therapy 
within 6 months of enrollment. 
f) Known Human Immunodeficiency Virus (HIV) infection or HIV-related disease. 
g) Hepatitis C virus infection. 
h) Genetic immunodeficiency. 
7. History of splenic or thymic dysfunction. 
8. Any serious chronic or progressive disease as assessed by the investigator (eg, neoplasm, 
hematologic malignancies, insulin dependent diabetes; cardiac, renal, or hepatic disease). 
9. Body Mass Index (BMI) greater than or equal to 35 kg/m2. 
10. Concurrent participation in any clinical trial with another investigational product 30 days prior 
to or during the conduct of this trial. 
11. Vaccination within 14 days (for inactivated vaccines) or 28 days (for live vaccines) prior to 
enrollment or plans to receive any vaccine within 28 days of trial vaccine administration 
(consider whether applicable as an exclusion criterion or criterion for delay of trial vaccine 
administration). 
12. Use of antipyretics and/or analgesic medications within 24 hours prior to vaccination. Trial 
entry should be delayed to allow for a full 24-hours to have passed since last use of 
antipyretics and/or analgesic medications (consider whether applicable as an exclusion 
criterion or criterion for delay of trial vaccine administration). 
13. Subjects with history of substance or alcohol abuse within the past 2 years. 
14. Subjects who are pregnant or breastfeeding. 
15. Subjects of childbearing potential who are sexually active with men and have not used 
“acceptable contraceptive methods” for at least 2 months prior to enrollment. 
a) Of “childbearing potential” is defined as beyond onset of menarche and not: menopausal 
for 2 or more years, post bilateral tubal ligation at 1 year prior, post bilateral 
oophorectomy for at least 1 year or post hysterectomy. 
b) “Acceptable birth control methods” include: 
a. Hormonal contraceptives (such as oral, injection, transdermal patch, 
implant, cervical ring). 
b. Barrier method (condom with spermicide or diaphragm with spermicide) 
every time during intercourse. 
c. Intrauterine device. 
d. Monogamous relationship with vasectomized partner (partner must have 
been vasectomized for at least 6 months prior to the subject’s enrollment. 

 
21 
16. Subjects of childbearing potential who are sexually active with men and refuse acceptable 
contraceptive method up to 28 days after the vaccination. 
17. Any positive or indeterminate pregnancy test. 
18. Planned vaccination (during the trial conduct) against any other vaccine preventable 
disease. 
19. Planned travel (during the trial) to any YFV endemic area. 
20. Screening serology consistent with prior history of dengue, zika, West Nile or Japanese 
encephalitis virus infection. 
21.  
 
It may occur that a prospective subject meets all entry criteria except one that relates to short 
term clinical condition (e.g., fever, recent use of excluded medications). Under these 
circumstances, eligibility for delayed trial enrollment may be considered after inclusion/exclusion 
criteria have been rechecked, and if the subject is confirmed to be eligible. 
 
8.3 Criteria for delay 
 
After enrollment, subjects may temporarily encounter circumstances that warrant delay in 
vaccination. If a subject meets a criterion for delay of vaccination, the subject may still be 
vaccinated after the window for delay has passed as long as the subject otherwise remains 
eligible for trial participation. 
 
The following clinical circumstances warrant a delay for administration of the vaccination: 
 
• 
Subjects with a clinically active infection (as assessed by the investigator) or body 
temperature ≥38°C (100.4°F) within 3 days of the vaccination date. 
• 
Subjects who have received, or have scheduled, any other vaccines within 30 days for 
both inactivated and live vaccines prior to trial vaccination. 
• 
Subjects who have used antipyretics and/or analgesic medications <=24 hours prior to 
vaccination. Trial vaccination should be delayed to allow for 24-hours to pass between 
having used antipyretics and/or analgesic medications and trial vaccine administration. 
 
8.4 Early Termination of subject trial participation  
Under certain circumstances, a subject’s trial participation may be terminated early. This means 
that no further trial procedures (including data collection) will be performed on that subject 
beyond the date of early termination of trial participation. The primary reason for early 
termination of the subject’s trial participation should be documented using the following 
categories. Although subject is not obligated to provide a reason for withdrawing, attempts 
should be made to determine the reason for the withdrawal and, where possible, the underlying 
reason should be documented. 
1. Adverse Event: The subject has experienced an AE (related/unrelated to the trial 
vaccination or trial-related procedures) that imposes an unacceptable risk to subject 
health and/or the subject is unwilling to continue participation because of the AE. In 
these cases, the primary reason for early termination of trial participation in this case will 
be ‘withdrawal due to AE’. Any ongoing AEs leading to early termination of trial 
participation should be followed up by the investigator until resolution or stabilization and 
safety follow-up will be offered. 
2. Lost to follow-up: The subject did not return to the trial site and at least 3 attempts to 
contact the subject failed. 

 
22 
3. Withdrawal of consent: The primary reason for early termination will be “withdrawal of 
consent” if the subject withdraws for a non-medical reason (i.e., reason other than AE). 
The reason for withdrawal, if provided, should be documented. 
4. Premature trial termination by a regulatory agency, the IEC/IRB, or any other authority. 
5. Subject’s death during trial participation. 
 
9 
TREATMENT OF SUBJECTS 
 
9.1 
Study Vaccine and Dosing Regimen 
YF-VAX® (YFV-17D) is manufactured and distributed by Sanofi Pasteur Inc. (Swiftwater, 
Pennsylvania). YF-VAX® is formulated to contain not less than 5.5 x 104 PFU/0.5 mL dose in 
single-dose or multiple-dose (5-dose) vials reconstituted with 0.6 mL or 3 mL of diluent, 
respectively (see YF-VAX® package insert for details). 
 
Storage 
YF-VAX® will be shipped frozen in a container containing solid carbon dioxide (CO2; dry ice) 
and will not be used if the shipping case contains no dry ice upon arrival. Upon receipt, 
lyophilized vaccine must be maintained continuously at 2˚C to 8˚C. The vaccine will not be used 
after the expiration date. The vaccine will not be re-frozen. YF-VAX® does not contain a 
preservative; all vaccine must be used within 1 hour after reconstitution. 
 
Vaccine Preparation 
 
The research pharmacist will reconstitute the vaccine per manufacturer’s instructions: YF-VAX® 
will be diluted to deliver not less than 5.5 x 104 PFU/0.5 mL dose in single-dose using the 
diluent supplied (sodium chloride for injection USP). The reconstituted vaccine should sit at 
room temperature for 1 to 2 minutes and then be carefully swirled until a uniform suspension is 
achieved. The reconstituted vaccine will appear slightly pink-brown in color. If the product 
contains extraneous particulate matter or is discolored, do not administer the vaccine. The 
vaccine will be swirled well and administered within 1 hour after reconstitution. 
 
Administration and Dosage 
 
Subcutaneous vaccination will deliver the standard 0.5 mL dose in the subject’s preferred 
deltoid. Tattoos and other skin findings that may obscure injection site reactions will be avoided. 
  
9.1.1 Interruption of Study Drug 
If unexpected AEs are noted, administration of YF-VAX® may be interrupted or discontinued.  
Dose modification is an unlikely course of action because the SC dose is the FDA-approved 
standard concentration.  
 
9.1.2 Vaccine Toxicity Management 
 
The clinical investigator will monitor all subjects for any vaccine-associated toxicity. 
The following plan has been developed to monitor subjects with severe or life-threatening 
vaccine-associated toxicities: 
 
• 
Study subjects will maintain a paper diary for local (injection site) for 7 days and 
systemic symptoms for 14 days post vaccination. Subjects will receive a thermometer 
and will record document temperatures at least once daily and more frequently if they 
experience a subjective fever. 

 
23 
• 
The study visit schedule ensures that subjects are seen by the investigator every other 
day +/- from post vaccination day 0 through 14. 
• 
At each visit, diary cards will be reviewed and a focused physical exam will be performed 
by the investigator as indicated by the schedule of events. 
• 
Liver function tests will be performed at post-vaccination days 0, 6, 14 and 28 to detect 
any abnormalities. 
• 
The investigator will be prepared to refer any subject requiring specialized treatment to a 
medical specialist (e.g., neurologist, gastroenterologist). 
 
9.2 
Documentation of Study Product 
The vaccine, YF-VAX®, will be administered at Oregon Health & Science University (OHSU), 
under the supervision of the PI. Careful logs will be kept to monitor receipt, dispensation, and 
destruction of drug through the pharmacy at each institution. The investigator is required to 
maintain adequate records of the disposition of the vaccine and placebo, including dates, 
quantity, and use by subjects as outlined in 21 Code of Federal Regulation (CFR) 312.62. If the 
investigation is terminated, suspended, discontinued, or completed, the investigator must 
destroy the unused supplies of the vaccine as required under 21 CFR 312.59. 
 
9.5 
Subject Compliance 
 
Compliance will be monitored by study personnel who will administer the vaccine. Subjects who 
miss a scheduled visit will be contacted by phone, text or email, and the visit will be re-
scheduled. 
  
10 
STUDY PROCEDURES 
 
10.1 Trial Procedures 
The following sections describe the trial procedures and data to be collected. All procedures 
must be performed by qualified and trained staff. 
 
10.1.1 Informed consent 
• 
Informed consent must be obtained prior to the subject entering into the trial, and before 
any protocol-directed procedures are performed. Non-English speaking participants will 
not be targeted but should the need arise to consent someone that is non-English 
speaking, a short form and interpreter services will be used. A modification will be 
submitted for approval of the needed short form. A unique subject number will be 
assigned to each subject after informed consent is obtained. If all eligibility criteria are 
fulfilled, this subject number will be used throughout the trial. Subject numbers assigned 
to subjects who fail screening should not be reused.  
10.1.2 Demographics, Medical History and Prior Medications 
• 
Demographic information will include age, sex, race, and ethnicity as described by the 
subject. 
• 
Medical history will also be collected, including but not limited to any medical history that 
may be relevant to subject eligibility for trial participation such as prior vaccinations, 
current medications, and previous and ongoing medical conditions. Relevant medical 
history may also include anything that contributes to the understanding of an AE that 
occurs during trial participation, if it represents an exacerbation of an underlying 
disease/preexisting problem. 
• 
Medical history will include any significant conditions or diseases that have disappeared 
or resolved at or prior to signing of the informed consent form. 

 
24 
• 
All medications, vaccines and blood products taken by the subjects are to be collected 
and documented in the subject’s source document: 
a) Medications: from 1 month (minimum 28 days) prior to vaccination and up to 1 month 
thereafter. 
b) Steroids and immunostimulants within 60 days prior to enrollment. 
c) Immunoglobulins and blood products within 3 months prior to enrollment. 
d) Immunosuppressive therapy within 6 months prior to enrollment. 
• 
The use of antipyretics and/or analgesic medications <= 24 hours prior to noted and the 
reason for their use documented in the subject’s source documents.  Vaccination should 
be delayed if subjects have used antipyretics and/or analgesic medications within 24 
hours of scheduled vaccine administration. 
• 
Prohibited therapies: 
a) Parenteral immunoglobulin, blood products, and/or blood-derived products within the 
3 months prior to enrollment. 
b) Immunosuppressive therapy within 6 months or systemic corticosteroid treatment 
within 60 days prior to enrollment, 
c) or immunostimulants within 60 days prior to enrollment. 
d) Any other vaccines within 30 days prior to vaccination and 30 days after vaccination. 
• 
Serological testing to ensure no prior Flavivirus infection. 
These data must be written in the subject’s source documents. 
10.1.3 Documentation of Trial Entrance/Randomization 
• 
Subjects who have a signed informed consent form, meet all of the inclusion criteria and 
none of the exclusion criteria are eligible for entrance into the active phase. 
• 
If the subject is found to be ineligible for enrollment, the investigator should record the 
primary reason for failure on the subject’s screening and enrolment log. 
• 
Randomization is not applicable for this open-label trial. 
10.1.4 Physical Examination 
• 
A complete physical examination must be performed by a qualified health professional in 
accordance with local regulations prior to vaccination. A complete physical examination 
includes but is not limited to: auscultation of heart and lungs, palpation of the abdomen, 
inspection of extremities (including skin over intended vaccination site), a check of 
general appearance and the measurement of weight and height; BMI will be calculated. 
Additional physical examinations may be performed if indicated by subject’s medical 
history. The findings should be documented in the subject’s source document. 
• 
Targeted symptom-directed physical examination including but not limited to 
measurement of vital signs may be performed on visit days 2, 4, 6, 8, 10, 12, and 14. 
Clinically significant changes, as determined by the investigator, from the baseline 
examination should be recorded in the subject’s source documents. 
10.1.5 Vital Signs 
• 
During the physical examination, a subject should have their vital signs measured, 
including (but not limited to) the measurement of systolic blood pressure/diastolic blood 
pressure, heart rate, and body temperature at all trial visits. 
10.1.6 Laboratory tests 
• 
Baseline complete blood count with differential and complete metabolic panel will be 
drawn on enrollment visit. 
10.1.7 Immunogenicity Assessments 
• 
Vaccine viremia will be measured using blood samples collected on post vaccination day 
0, 2, 4, 6, 8, 10, 12, and 14. 
• 
Yellow fever neutralizing antibodies will be measured using blood samples collected 
from all subjects prior to first vaccination on day 0 and day 28. 

 
25 
• 
T and B cell-mediated immune response will be measured using blood samples 
collected from all subjects collected post vaccination day 00, 2, 8, 14, and 28. 
• 
The innate immune response will be measured using blood samples collected post-
vaccination days 0, 2, 8, 14 and 28. 
• 
All blood samples will be processed, labeled and stored according to the Laboratory 
Manual or other appropriate guideline provided to the trial site. 
10.1.8 Processing, Labeling and Storage of Biological Samples 
• 
PBMC will be collected, processed, labeled and securely stored according to laboratory 
Standard Operating Procedures (SOP). 
• 
All biological samples will be processed, labeled and stored according to SOP or other 
appropriate guideline provided to the site. 
10.1.9 Safety Assessments 
• 
Safety assessments will include collection and recording of solicited local (injection site) 
reactions and solicited systemic events, unsolicited AEs (serious and non-serious), and  
10.1.10 
Clinical Safety Laboratory Variables 
• 
Liver function tests will be performed on post vaccination days 0, 6, 14, and 28 
10.1.11 
Contraception and Pregnancy Avoidance Procedure 
• 
For female subjects of child bearing potential, point of care urine pregnancy testing will 
be performed prior to vaccination (Day 0). Results must be confirmed and documented 
as negative prior to vaccination. Subjects will be provided with information on acceptable 
methods of contraception as part of the subject informed consent process and be asked 
to sign a consent form stating that they understand the requirements for avoidance of 
pregnancy and donation of ova. 
10.2 Monitoring Subject Compliance 
The investigator must record all injections of trial vaccine given to the subject in the subject’s 
source document. 
10.3 Schedule of Observations and Procedures 
10.3.2 Pre-Vaccination Procedures 
1. Before performing any trial procedure, the signed informed consent form needs to be 
obtained. 
2. Check inclusion and exclusion criteria. 
3. Review demographic data, medical history, and prior medication/vaccination. 
4. Review of systems: a structured interview that queries any complaints the subject has 
experienced across each organ system. 
5. Perform a complete physical examination. 
6. Check vital signs.  
7. Perform pregnancy testing (serum or urine) for females of childbearing potential.  
8. Provide avoidance of pregnancy guidance for females of childbearing potential who are 
sexually active. 
9. Collect pre-vaccination blood samples 
10.3.3 Vaccination Procedures 
1. Check criteria for delay of trial vaccine administration.  
2. Check contraindications for trial vaccine administration.  
3. Administer the trial vaccine.  
10.3.4 Post Vaccination Procedures 
1. Review with the subject how to measure solicited local (injection site) reactions and body 
temperature, how to complete the diary card and how often to complete the diary card. 
Training of the subject on how to measure an injection site reaction and how to take their 
temperature, as well as how to record the information in the diary card, should be 
performed while the subject is under observation after vaccination. 

 
26 
2. The subject must understand that timely, daily completion of the diary card is a critical 
component of trial participation. Subjects should be instructed to write clearly and to 
complete the diary card in pen. Any corrections to the diary card by the subject should 
include a single strikethrough line with a brief explanation for any change and be initialed 
and dated. Diary cards will be the only source document for remote collection of solicited 
local (injection site) reactions, solicited systemic events (including body temperature 
measurements), and unsolicited (non-serious) AEs.  
3. The diary card should be reviewed with the subject. 
4. No corrections or additions to the diary card will be allowed after it is reviewed by the 
investigator. 
5. Any data that is identified as implausible or incorrect, and confirmed by the subject to be 
a transcription error, should be corrected by the subject on the diary card with a single 
strikethrough line and should be initialed and dated by the subject. 
6. Any newly described solicited and unsolicited (non-serious) safety information should be 
added to the diary card by the subject, initialed, and dated. 
7. Starting on the day of vaccination, the subject will check for specific types of events at 
the injection site (solicited local [injection site] reactions), any generalized symptoms 
(solicited systemic events) including body temperature, unsolicited (non-serious) AEs, 
any other change in the subject’s health status, and any medications taken. Solicited 
local (injection site]) reactions, solicited systemic events, body temperature, and 
unsolicited (non-serious) AEs will be recorded in the diary. Assessments should 
preferably take place at the end of the day. 
8. Body temperature measurement is to be performed using a thermometer provided by the 
study team. Temperatures should be taken at approximately the same time of day, 
preferably at the end of the day. If the subject feels unusually hot or cold during the day, 
the subject should check his/her temperature. If the subject has fever, the highest body 
temperature observed that day should be recorded on the diary card. 
9. The measurements of solicited local (injection site) reactions are to be performed using 
the ruler provided by the study team. 
10. The collection on the diary card of solicited local (injection site) reactions, solicited 
systemic events (including body temperature measurement), and unsolicited (non-
serious) AEs will continue for a total of 7 days, 14 days, and 28 days following 
vaccination. Any solicited local (injection site) or systemic AE that resolves before 8 or 
15 days, respectively, following each trial vaccination, but recurs at a later time (ie, if 
discontinues), should be recorded as an unsolicited AE on the “Adverse Event”. 
11. After each trial vaccination, the subject will be observed for at least 30 minutes including 
observation for solicited local (injection site) reactions, solicited systemic events 
(including body temperature measurement), and unsolicited AEs. The investigator or 
delegate will take the opportunity to remind the subject how to measure solicited local 
(injection site) reactions and body temperature as part of this observation period. All 
safety data will be collected in the subject’s source documents. 
12. The site should schedule the next trial visit with the subject. 
13. The subject will receive a written reminder of the next trial visit. 
14. The subject will be reminded to complete the diary card daily, to contact the site if there 
are any questions, and to contact the site immediately (or as soon as the subject is 
medically stable) if the subject has a medical condition that leads to a hospitalization or 
an emergency room visit or is otherwise perceived as serious. All contact details will be 
provided to the subject. 
10.3.5 Site Visits after Vaccination: Days 2, 4, 6, 8, 10, 12, 14, 28. 
1. Review the diary card with the subject days 2, 4, 6, 8, 10, 12, 14. 
2. Collect and record persistent/prolonged solicited local (injection site) reactions. 

 
27 
3. Collect and record persistent/prolonged solicited systemic events. 
4. Collect concomitant medications/vaccinations. 
5. Perform a complete physical examination on day 28. 
6. Check vital signs.  
7. Provide guidance with respect to the avoidance of pregnancy for females of childbearing 
potential who are sexually active. 
8. Collect and report SAEs.  
9. Collect blood samples. Blood should be taken from the subject using an aseptic 
venipuncture technique. 
10. The investigator or delegate should schedule the next trial visit with the subject, as 
applicable. 
11. The subject will receive a written reminder of the next trial visit, as applicable. 
12. The subject will be reminded to complete the diary card daily and to contact the site if 
there are any questions, and to contact the site immediately if the subject has a medical 
condition that leads to a hospitalization or an emergency room visit or is otherwise 
perceived as serious. All contact details will be provided to the subject. 
10.3.6 Final (End of Trial) Visit 
The final (end of trial) visit will be performed on Day 28. If a subject terminates earlier, the final 
(end of trial) visit procedures should be performed at their last trial visit, if possible. 
 
10.3.7 Post-Trial Care 
No post-trial care will be provided. 
 
10.4 
Sample Collection 
 
10.4.1 Blood Samples 
 
Primary and secondary endpoint assessments and clinical safety labs will be performed using 
blood samples collected at each study visit. All blood samples will be processed, labeled and 
stored according to the appropriate guideline provided to the site. Samples will be preserved 
and retained at a central laboratory indefinitely or as required by applicable law. The 
Investigator has put into place a system to protect the subject’s personal information to ensure 
optimal confidentiality and defined standard processes for sample and data collection, storage, 
analysis, and destruction. 
 
10.4.2 PBMC Cryopreservation 
 
The blood samples obtained on post vaccination days 0, 2, 8, 14 and 28 will be a source of 
PBMCs and serum for studies involving innate and cellular immune responses to viral infection. 
PBMCs from heparinized whole blood will be collected and cell numbers will be determined. 
Multiple aliquots will be appropriately labeled with a unique identification code, cryopreserved, 
and stored in liquid nitrogen. 
 
11 
SAFETY ASSESSMENTS 
 
11.1 
Safety Assessments Overview 
 
Adverse events that are classified as serious (see Section 11.2.3) according to the definition of 
21 CFR 312.32 must be reported promptly and appropriately to institutional review boards 
(IRBs) and health authorities. This section defines the types of AEs and outlines the procedures 
for appropriately collecting, grading, recording, and reporting them. 

 
28 
 
Information in this section complies with ICH Guideline E2A: Clinical Safety Data Management: 
Definitions and Standards for Expedited Reporting, ICH Guideline E-6: Good Clinical Practice 
(GCP) and applies the standards set forth in the Toxicity Grading Scale for Healthy Adult and 
Adolescent Volunteers Enrolled in Preventive Vaccine Clinical Trials guidelines (September 
2007)(33). This document provides a common vocabulary for describing grades of severity to 
analyze and interpret data, and for articulating the clinical significance of all AEs. The Toxicity 
Grading Scale for Healthy Adult and Adolescent Volunteers Enrolled in Preventive Vaccine 
Clinical Trials guidelines (September 2007) have been reviewed by the PI and have been 
deemed appropriate for the subject population to be studied in this protocol. 
 
Subjects will be asked about signs and symptoms at each clinical encounter. Clinical laboratory 
data will be analyzed to address safety issues. Physical examinations will be performed and 
vital signs collected (including: heart rate, respiration rate, body temperature, blood pressure). 
 
11.2 
Definitions 
 
11.2.1 Adverse Events 
 
An AE is any occurrence or worsening of an undesirable or unintended sign (including an 
abnormal laboratory finding), symptom, or disease that is temporally associated with the use of 
a study product, whether considered related to the study product or not. 
 
Expected AEs: Adverse events associated with YF-VAX® may include redness, pain, and 
swelling at the injection site, mild headaches, myalgia, low-grade fevers, or other minor 
symptoms for 5 to 10 days post-vaccination. Local reactions — including edema, 
hypersensitivity, pain, or mass at the injection site — have also been reported following YF 
vaccine administration. Immediate hypersensitivity reactions — characterized by rash, urticaria, 
and/or asthma — are uncommon and occur principally among persons with a history of egg 
allergy. Approximately 72% of subjects experienced non-serious AEs judged to be related to 
vaccination. Most of these were injection site reactions of mild to moderate severity. 
 
Solicited local (injection site) reactions and systemic AEs 
• 
Local reactions 
o Pain 
o Erythema 
o Swelling 
• 
Systemic events 
o Fever 
o Headache 
o Asthenia 
o Malaise 
o Myalgia 
o Rash 
Severity scale for solicited Safety Parameters 
• 
Pain at injection site 
0 None 
1 Mild – no interference with activities of daily living (ADLs) 
2 Moderate – interference with ADLs without treatment 
3 Severe – prevents ADLs without treatment 

 
29 
• 
Erythema at injection site 
0 <25 mm 
1 Mild: >25-<50 mm 
2 Moderate: >50-<100 mm 
3 Severe: >100mm 
• 
Swelling at injection site 
0 <25 mm 
1 Mild: >25-<50 mm 
2 Moderate: >50-<100 mm 
3 Severe: >100mm 
• 
Headache 
0 None 
1 Mild – no interference with activities of daily living (ADLs) 
2 Moderate – interference with ADLs without treatment 
3 Severe – prevents ADLs without treatment 
• 
Asthenia 
0 None 
1 Mild – no interference with activities of daily living (ADLs) 
2 Moderate – interference with ADLs without treatment 
3 Severe – prevents ADLs without treatment 
• 
Malaise 
0 None 
1 Mild – no interference with activities of daily living (ADLs) 
2 Moderate – interference with ADLs without treatment 
3 Severe – prevents ADLs without treatment 
• 
Myalgia 
0 None 
1 Mild – no interference with activities of daily living (ADLs) 
2 Moderate – interference with ADLs without treatment 
3 Severe – prevents ADLs without treatment 
 
 
11.2.2 Medically Attended Adverse Events 
 
MAEEs are defined as AEs that result in an unscheduled medical visit by a healthcare 
professional, including visits to an emergency department, but not fulfilling seriousness criteria. 
 
11.2.3 Serious Adverse Events 
 
An SAE or reaction is defined as “any AE occurring at any dose that suggests a significant 
hazard, contraindication, side effect, or precaution.” This includes, but is not limited to, any of 
the following events: 
1) 
Death: A death that occurs during the study or that comes to the attention of the 
investigator during the protocol-defined follow-up period after the completion of therapy must be 
reported to the independent safety monitor (ISM) whether it is considered treatment related or 
not. 
2) 
A life-threatening event: A life-threatening event is any adverse therapy experience that, 
in the view of the investigator, places the subject at immediate risk of death from the reaction as 
it occurred. 
3) 
Inpatient hospitalization or prolongation of existing hospitalization 
 

 
30 
4) 
Persistent or significant disability/incapacity 
 
5) 
Congenital anomaly or birth defect 
 
An important medical event that may not result in death, be life-threatening, or require 
hospitalization may be considered an SAE when, based on appropriate medical judgment, it 
may jeopardize the subject and may require medical or surgical intervention to prevent one of 
the outcomes listed above. 
 
Serious AE reporting will begin with events that arise after the informed consent is signed until 
30 days after the last study visit. 
 
11.2.4 Unexpected Adverse Events 
 
An AE is considered “unexpected”, when the nature (specificity) or severity of the AE is not 
consistent with applicable product information, such as safety information provided in the 
package insert. 
 
11.3 
Collection and Recording of Adverse Events 
 
11.3.1 Collection Period 
 
Adverse events will be collected from the time the subject signs the study consent until 30 days 
after he/she completes study participation, or until 30 days after he/she prematurely withdraws 
from the study. 
 
Once recorded, an AE will be followed until it resolves with or without sequelae, becomes 
medically stable, or until 30 days after a participant completes or terminates from the study, 
whichever comes first. Where indicated, particularly in the event of neurological, 
gastrointestinal, or cardiac problems, the subject will be referred to appropriate medical 
specialists. 
 
11.3.2 Collection of Adverse Events 
 
Adverse events (including SAEs) may be discovered through any of these methods: 
• 
Observing the subject 
• 
Questioning the subject in an objective manner 
• 
Receiving an unsolicited complaint from the subject 
• 
Reviewing subject diary information 
 
In addition, an abnormal value or result from a clinical or laboratory evaluation (e.g., a 
radiograph, an ultrasound, or an electrocardiogram) can also indicate an AE. If this is the case, 
then the evaluation that produced the value or result should be repeated until that value or result 
returns to baseline or can be explained and the subject’s safety is not at risk. If an abnormal 
value or result is determined by the investigator to be clinically significant, it must be recorded 
as an AE. 
 
11.3.3 Recording Adverse Events 
 
Throughout the study, the investigator will record and grade AEs using the appropriate source 
document and will enter it into the system regardless of event severity or relation to study 

 
31 
product or study procedure. All requested information on the source document should be 
provided and entered, if available. 
 
For a fatal outcome, cause of death and a comment on its possible relationship to the study 
treatment should be provided. For each AE or SAE, one of the following outcomes will be 
determined: 
• 
Ongoing stable 
• 
Ongoing unstable 
• 
Resolved without sequelae 
• 
Resolved with sequelae (this could also be stability) 
• 
Death 
 
Actions taken in response to an AE and follow-up results (including lab results) will also be 
recorded in the subject’s medical record in accordance with local procedure. Any treatment 
administered for the AE must be recorded in the subject’s EDC source document and entered 
into the EDC system. When a subject is discontinued from the study due to an AE, relevant 
clinical assessments and laboratory tests will be repeated as necessary until final resolution, 
stabilization, or until 30 days after discontinuation occurs, whichever comes first. 
 
11.3.4 Recording Serious Adverse Events 
 
Serious AEs will be recorded in REDCap, the EDC, on the Serious Adverse Event Report Form, 
which will then be sent to the medical monitor about the SAE (Section 11.5.2). In addition, SAEs 
will be recorded in the EDC source document and entered into the EDC system. All requested 
information on the AE EDC form should be provided, if available. 
 
11.4 
Grading and Attribution of Adverse Events 
 
11.4.1 Grading Criteria 
 
The study site will grade the severity of AEs experienced by the study subjects according to the 
criteria set forth in the Toxicity Grading Scale for Healthy Adult and Adolescent Volunteers 
Enrolled in Preventive Vaccine Clinical Trials guidelines: 
 
Adverse events will be graded on a scale from 1 to 4 according to the following standards in the 
Toxicity Grading Scale for Healthy Adult and Adolescent Volunteers Enrolled in Preventive 
Vaccine Clinical Trials guidelines: 
Grade 1 = mild AE.  
Grade 2 = moderate AE. 
Grade 3 = severe and undesirable AE. 
Grade 4 = life-threatening AE, disabling AE, or death. 
 
11.4.2 Attribution Definitions 
 
The relation, or attribution, of an AE to a vaccine will be determined by the site investigator. The 
site investigator will also record the determination of attribution on the appropriate EDC source 
document, entered into the EDC system, and/or SAE reporting form. The relation of an AE to 
the vaccination or study procedures will be determined using the descriptors and definitions 
provided in Table 11.1. 
  
 

 
32 
Table 11.1 
Toxicity Grading Scale for Healthy Adult and Adolescent Volunteers Enrolled in 
Preventive Vaccine Clinical Trials Guidelines Attribution of Adverse Events. 
 
Code Descriptor 
Definition 
Unrelated Category 
1 
Unrelated 
The AE is clearly not related to the vaccination. 
Related Categories 
2 
Unlikely 
The AE is doubtfully related to the vaccination. 
3 
Possible 
The AE may be related to the vaccination. 
4 
Probable 
The AE is likely related to the vaccination. 
5 
Definite 
The AE is clearly related to the vaccination. 
 
For additional information and a printable version of the Toxicity Grading Scale for Healthy Adult 
and Adolescent Volunteers Enrolled in Preventive Vaccine Clinical Trials guidelines, consult the 
FDA website: http://fda.gov/cber/gdlns/toxvac.htm. 
 
11.5 
Reporting of Adverse Events 
 
11.5.1 Adverse Events Requiring Expedited Reporting 
 
The following AEs, as graded with the Toxicity Grading Scale for Healthy Adult and Adolescent 
Volunteers Enrolled in Preventive Vaccine Clinical Trials guidelines (Section 11.4.1) will be 
reported in an expedited fashion by investigators to the institutional review board of receipt 
(Section 11.5.2), regardless of expectedness or relationship to the vaccination: 
• 
All SAEs per 21 CFR 312.32 definitions (Section 11.2.3) 
• 
All Toxicity Grading Scale for Healthy Adult and Adolescent Volunteers Enrolled in 
Preventive Vaccine Clinical Trials guidelines Grade 3 and 4 (33) 
• 
Any event that the investigator considers serious but is not easily categorized 
  
11.5.2 Reporting Timeline 
 
The following process for reporting an SAE ensures compliance with ICH guidelines. When an 
investigator identifies an SAE (as defined in Section 11.2.3) or other AE requiring expedited 
reporting (see Section 11.5.1), the ISM must be notified immediately upon discovery of the 
event, and must complete and send the Serious Adverse Event Form within 1 business day to 
the Independent Safety Monitor within 2 business days after receipt of the SAE report from the 
site. 
 
All non-serious AEs (defined in Section 11.2.1) will be recorded on the appropriate EDC source 
document and entered into the AE form included in the EDC system. Non-serious AEs will be 
reported based on the guidelines of the institutional review board. 
 
The site investigator will follow the progress of a subject who experiences a SAE until the SAE 
is resolved, considered stable, or until 30 days after the subject completes or terminates from 
the study, whichever comes first. If the unexpected SAE has not resolved by the report 
deadline, the investigator will send follow-up reports. 
 
11.5.3 Reporting of Adverse Events  
 
All AEs (serious or otherwise) All SAEs will be reported per YFVax package insert, "To report 
SUSPECTED ADVERSE REACTIONS, contact the Pharmacovigilance Department, Sanofi 

 
33 
Pasteur Inc., Discovery Drive, Swiftwater, PA 18370 at 1-800-822-2463 (1-800VACCINE) or 
VAERS at 1-800-822-7967 or https://vaers.hhs.gov.  
 
11.5.4 Reporting Pregnancy as a Serious Adverse Event 
 
This study requires that any pregnancy be reported as an SAE manufacturer for tracking 
purposes only. All pregnancies identified during the study must be followed to conclusion and 
the outcome must be reported. The investigator should be informed immediately of any 
pregnancy in a study subject. The investigator should report all pregnancies to the ISM within 1 
business day (see Section 11.5.2). The investigator should counsel the subject and discuss the 
risks of continuing with the pregnancy and the possible effects on the fetus. Monitoring of the 
pregnant subject should continue until the conclusion of the pregnancy, and a follow-up SAE 
reporting form detailing the outcome of the pregnancy should be submitted. 
 
Information requested about the delivery will include: 
 
• 
Subject’s enrollment identification 
• 
Gestational age at delivery 
• 
Birth weight, length, and head circumference 
• 
Gender 
• 
Appearance, pulse, grimace, activity, and respiration (APGAR) score at 1 minute, 5 
minutes, and 24 hours after birth, if available 
• 
Any congenital abnormalities 
 
11.6 
Review of Safety Information 
 
11.6.1 Routine ISM Review 
 
The ISM will generate monthly reports that compile all newly submitted and accumulated AEs 
and SAEs. Subsequent reviews of the periodic reports will be performed by the FDA. 
 
11.6.2 ISM Product Safety Associate Review 
 
The ISM will review all SAEs immediately upon notification by the Investigator. These reports 
will consist of all AEs and SAEs. 
 
11.6.3 Independent safety monitor. An independent safety monitor will be identified as 
mandated by the OHSU IRB in data safety and monitoring plan (DSMP). 
 
11.6.4 Institutional Review Board Reporting of Adverse Events 
 
The investigator must report AEs to OHSU IRB as locally mandated.  
 
11.7 
Protocol Deviations 
 
The study coordinator will report all protocol deviations within 1 business day by completing the 
EDC Protocol Deviation Form. This form will ask for a description of the event and what 
corrective action is planned. All protocol deviations that meet reporting criteria will be reported to 
the OHSU IRB within 5 business days. 
 

 
34 
Protocol deviations occur under the following circumstances: when non-adherence to the 
protocol results in a significant added risk to the study subject; when the study subject or 
investigator has failed to adhere to significant protocol requirements; or when there is non-
adherence to GCP guidelines.  NIAID will not allow waivers or planned deviations of enrollment 
criteria; enrollment of subjects who do not meet the inclusion criteria constitutes a deviation from 
the protocol. 
 
11.8 
Clinical Monitoring 
 
Study staff unrelated to any other aspect of the trial will monitor the study to verify that the rights 
and well-being of the subjects enrolled under this protocol are protected and that the data 
being collected by the site are accurate, complete, and verifiable from source documents. The 
study PI will verify that the site complies with the currently approved protocol and any protocol 
amendments, and with GCP, and any other regulatory requirements. The site will be monitored 
in accordance with the monitoring plan developed for this clinical study. 
 
12 
DATA HANDLING AND RECORD KEEPING 
 
12.1 
EDC and Source Documentation 
 
The site study coordinator will complete an EDC source document for each subject. Completion 
instructions for the EDC source document are embedded in the document and an online help 
function is provided to assist with EDC entry. Information in the EDC system must be currently 
maintained and up-to-date. 
 
Subjects must not be identified by name on any study document. Subjects will be identified by 
subject identification numbers assigned at enrollment. 
 
12.2 
Data Management 
 
Data for this project will be collected and stored in OCTRI's installation of REDCap, a highly 
secure and robust web-based research data collection and management system. Using this 
system, clinical site personnel use an internet browser to key data into electronic data fields. 
Univariate data validation tests will be performed as the data are keyed, and most implausible 
data values will be resolved immediately. Authorized site personnel may log in to the system at 
any time, review and correct previously entered data, and key additional data. 
 
Adverse events will be coded as detailed in the safety monitoring plan. 
 
Upon completion of processing, the operational database will be subjected to database closure 
procedures and subsequently locked. The PI will sign-off on eCRFs after review. 
 
After the operational database is locked, the programming team will create and validate an 
analysis database. The analysis database will be the basis for statistical analyses of the data. 
 
Physical security: The KPNW CHR and OHSU OCTRI laboratories (location of specimen 
storage freezers) and the offices of research personnel have limited access.  
Additionally, the PI’s and research staff’s office doors are kept locked when they are not 
there. The OHSU OCTRI and Messer Laboratories are in locked rooms with access 
limited to research personnel. OCTRI's REDCap software is housed on servers located 
in ITG's Advanced Computing Center providing locked physical security. 

 
35 
Electronic security: Associated data will be held securely in compliance with OHSU and 
KPNW Information and Security Guide.  Specifically, data will be held on OHSU 
networked drives, behind the OHSU firewall.  Files will be password-protected. The 
REDCap servers are housed behind both the OHSU firewall and a second ACC firewall.  
All web-based data transmissions are encrypted with industry-standard SSL methods.   
Controlled User Access: REDCap employs a robust multi-level security system that 
enables researchers to easily implement "minimum necessary" data access for their 
research staff, including specification of data fields that are identifiers. This feature 
includes “single click” ability to provide completely deidentified (removing all identified 
data fields and shifting dates) for analysis or other purposes.  User activities are logged 
to enable auditing of all data access.   Access is integrated with OHSU's network such 
that users who are also OHSU employees are authenticated against their OHSU 
network credentials. 
Data Integrity: REDCap is jointly managed in accordance with OHSU Information 
Security Directives by ACC staff and members of OCTRI's Biomedical Informatics 
Program, ensuring fidelity of database configuration and back-ups.  User activities are 
logged to enable auditing of all data changes. 
 
12.3 
Data Monitoring 
 
After study initiation, clinical research associates will monitor EDC system data and source 
documents, as well as, compliance with the protocol, and subject safety. 
 
12.4 
Record Retention 
 
The study-related records will be maintained at the site using either OHSU’s electronic medical 
record, EPIC, or Florence eBinders, which is a 21 CFR Part 11 compliant database to be used 
for clinical trial research. These records will include the following: 
1) 
Copies of the protocol and all amendments 
2) 
Signed Investigator Page for the protocol and all amendments 
3) 
 
4) 
Approval letter(s) from, and all other correspondence to and from, the IRB 
5) 
Curricula vitae (CVs)(current within 2 years at study initiation) and current medical 
licenses for the PI and any sub-investigators 
6) 
Delegation log of all study personnel 
7) 
The original signed informed consent form for each subject screened and signed 
consent forms  
8) 
Copies of results of all laboratory tests and other original data from which CRF 
information was obtained, as well as, laboratory normal ranges and certifications, when 
indicated 
9) 
Copies of all correspondence relating to this investigation 
10) 
Copies of financial disclosure documents 
 
13 
STATISTICAL ANALYSIS 
 
13.1 
Analysis Populations 
 
13.1.1 Analysis Population 
 

 
36 
All eligible, enrolled, and randomized subjects who have both a baseline antibody titer (obtained 
during the Screening Visit) and a Day 28 titer will be included in the Analysis population. 
 
13.2 
Statistical Methods 
 
13.2.1 Demographics, Baseline Characteristics, and Disposition 
 
Background and baseline demographic, physical examination, and subject disposition data will 
be tabulated and presented for all enrolled subjects.  Disposition will be tabulated according to 
the number (%) of subjects who have: signed informed consent, provided baseline/screening 
information, provided baseline blood draw for YF titer, completed each of the follow-up study 
visits for blood draw, and completed Day 28 (Visit 8) study procedures. Continuous data (e.g., 
age, body weight, height) will be summarized descriptively by mean, standard deviation (SD), 
median, and range.  Categorical data (e.g., gender, race) will be presented as enumerations 
and percentages. 
 
13.2.2 Primary Analysis –  
Fold change between pre and post-vaccination neutralizing antibody titer. Subject serum is 
serially diluted four-fold from starting dilutions of 1:5 and mixed with an equal volume of a fixed 
number of plaque or focus forming units (FFU) added to 96 wells plates seeded with Vero cells 
24 hours prior, and overlaid with methylcellulose. After 3 days incubation overlay is removed, 
cells are fixed and immune-stained, visualized, and counted on a CTL Immunospot™ S6Macro 
automated plate reader. Experiments will be conducted in 3 biologic replicates to ensure stable 
titers estimates. Foci counts are quality controlled by a technician blinded to sample exposure 
history. Positive and negative control sera will be run with each set of unknown sera. Foci 
counts will be transformed into percent neutralization by serum dilution and NT90 values will be 
calculated by sigmoid dose-response curve fitting. 
 
13.2.3 Secondary Analyses 
 
Viral load. Quantitative reverse-transcription PCR will be used to quantify viral genomic 
copies/mL in the vaccine inoculum and acute (Days 2-14) post-vaccination sera as previously 
described46,63 modified for 96-well high-throughput using the Qiacube™ sample preparation 
robot in the OHSU Gene Profiling Shared Resource (GPSR) for RNA extraction quantified using 
the QuantStudio ™ 12K Flex instrument supported by the GPSR. Virus quantification will also 
be done simultaneously on both vaccine inoculum and acute sera by direct plaque assay as 
previously described55, modified for 96-well plates, and virus isolation in tissue culture. To 
ensure rigor and reproducibility, all technicians will be blinded to subject study ID, known copy 
number positive controls and virus controls will be run with each batch. 
 
Area under the curve receiver operator characteristic (AUC ROC). We will conduct AUC ROC 
cutoff analyses to identify post-hoc the most sensitive and specific cutoff NT90 serum dilution 
threshold to predict post-vaccination boost and, if numbers are sufficient, for viral load as well.  
 
Neutralization titer AUC ROC. While 90% neutralization (NT90) is the most widely used NT 
percentage for evaluating YFV vaccinee neutralizing Ab titers, there are no data-based 
justifications for the selection of 90% neutralization in the assay. Alternatives include 50% 
neutralization, and NT50 is commonly used in the DENV field, less frequently in YFV field (e.g. 
22,32). As with NT90, no data support NT50 as the best evaluation point for NT assays. However, 
Salje et al.76 analyzed of over 2,000 repeated DENV PRNT done at WRAIR and found that 75% 
(NT75) was the optimal evaluation point for DENV NT assays based on square bias, variance, 

 
37 
and mean square error of NT estimates compared over a range of NT40-NT90. In Aim 3, we 
evaluate the clinically meaningful outcome of sterile protection from live attenuated virus 
challenge. Here we will use AUC ROC analysis to evaluate alternate NT evaluation points (NT75 
etc.) and identify the NT evaluation point that best discriminates (sensitive and specific) the NT 
protective threshold against boost and/or viral load. Using this clinically meaningful NT 
evaluation point we will re-assess our primary outcome in Aim 3 which is based on an NT90 
threshold of >1:40 and similarly re-evaluate NT threshold-based tests in Aims 1 and 2.  
 
13.2.4 Safety Data 
 
Safety-related outcomes (Section 11) will be monitored throughout the study period. All AEs 
occurring on or after YF vaccination will be summarized by body system. If a subject 
experiences more than one AE, the subject will be counted only once for that AE at the greatest 
intensity or relationship to YF vaccination. Adverse events will also be summarized according to 
seriousness, severity, and relationship to the vaccination. 
 
13.2.5 Safety Review 
 
Analyses of safety data will be conducted by the ISM after the first three subjects and at two 
month interim time points during the study. 
 
Additional safety reviews will occur whenever the stopping rules are met as specified in Section 
7.3. 
 
13.3 
Sample Size 
 
To execute this study we will work with Dr. Naleway at KPNW to identify previously vaccinated 
subjects 8 or more years post vaccination. KNPNW vaccinates ~400 individuals/year aged 20-
49 years old. Assuming we contact 400 subjects/year with ~4% uptake, we expect to recruit 
seventeen subjects/year for 2 years for a total of 34 subjects over the study period. Subjects will 
be recruited pre-vaccination and followed for 28 days post-vaccination with blood draws on days 
01 2, 4, 6, 8, 10, 12 ,14 and 28. We assume little to no loss-to-follow up as this study will be 
conducted over only 65 days, at the end of study Year 5 we expect we will have at least 32 
subjects to fully evaluate. 
 
Boost Power calculations. Based the combined studies of Reinhardt27, Hepburn31 and 
Kongsgaard32, we will assume a priori a sterilizing threshold of NT90 >1:40. Hepburn provides a 
robust estimate of 50% (500/1029) of subjects with NT80 <1:40 at 10 years post-vaccination31, 
with the caveat that some of these subjects may have been boosted previously. Assuming that 
10% of 16 subjects with titers >1:40 boost, and 75% of 16 subjects with titers <1:40 boost, 
against the null that 50% will boost each in group, the study has power=0.85 (χ2=12.28, DF=3, 
α=0.05, 32 subjects) to detect a difference in boost between two groups divided at an NT90 
threshold of >1:40. The study retains power >0.80 with as few as 22 subjects under the same 
assumptions, and 20 subjects will have power=0.76. There are no prior data available to 
estimate the proportions of subjects who will develop a detectable viral load on 17D re-
vaccination, and we consider evaluation for viral load exploratory but extremely important for the 
field. We assume that viral load is most likely to be detected in re-vaccinated subjects with 
NT90<1:40, or 16 (50%) subjects, and we contend that the null hypothesis is that no more than 1 
in 100 will have a detectable viral load as this is consistent with reasoning behind CDC and 
WHO recommendations for once-in-a-lifetime vaccination. Under these assumptions, the study 
has power=0.80 (test of 1 proportion75) to detect a viral load in at least 1 in 10 subjects. 

 
38 
 
AUC ROC power calculations. Assuming 18 of 32 subjects boost, the study will have a power 
=0.80 (α=0.05)66 to detect AUC >0.77 (fair to good range), similar to the AUC=0.740 reported by 
Kongsgaard et al.32 for the same analysis with 28 subjects. In order for viral load to be 
informative at the same level, at least 13 subjects would need to have detectable viral loads.  
 
14 
HUMAN SUBJECTS PROTECTION 
 
14.1 
Discontinuation of Study 
 
The OHSU IRB reserve the right to discontinue the study at any time for administrative reasons. 
Should either party discontinue the study prior to completion, the subjects already enrolled in 
the study will be allowed to complete the study, if medically appropriate. In such an event, each 
enrolled subject will be followed through the period outlined in the study. Investigators will be 
reimbursed for reasonable expenses incurred to the date of discontinuation on the basis of 
completed subjects. 
 
14.2 
Ethics 
 
14.2.1 Good Clinical Practice and Institutional Review Board Review 
 
Compliance with GCP guidelines for the conduct and monitoring of this clinical study will occur 
through observation of the ethical and regulatory requirements presented in ICH E6, Good 
Clinical Practice: Consolidated Guideline. By signing this protocol, the investigator agrees to 
adhere to these requirements. The study (protocol, informed consent, advertisements, product 
insert, subject information sheets, and investigator CV and credentials) must be reviewed and 
approved by the IRB. Changes to study materials (protocol, informed consent and 
advertisements) must be approved by the local IRB. 
 
14.2.2 Informed Consent 
 
Informed consent is an ongoing process that includes the signing of an informed consent form.  
Subjects are required to sign an informed consent form prior to being screened, and before 
undergoing any study procedures or assessments.  When substantial modifications are made to 
the informed consent form, the local IRB may require that all subjects currently enrolled in the 
study be re-consented.  Subjects will be provided with a copy of the signed informed consent 
form and printed materials that explain the purpose of the study, the medication(s) used in the 
study, procedures, and assessments. Subjects will also be provided with the telephone numbers 
of the investigator and qualified personnel who can assist with their questions and concerns. 
 
14.3 
Study Risks 
 
14.3.1 Vaccine Risks 
 
YFV-17D is Food and Drug Administration (FDA)-approved in the US for both primary 
vaccination in persons aged >6 months traveling to YFV endemic regions and booster 
vaccination in subjects considered high risk for YFV exposure. 17D has a strong record of 
safety and tolerability dating back to its introduction in 1936. Safety data suggest that booster 
vaccination as at least 10-fold lower risk of a vaccine related severe adverse event (SAE) 
compared to primary vaccination. The CDC reports data74 from observational studies that 
include 333 million vaccine doses, with 1,255 SAEs overall; vaccination type (primary vs boost) 

 
39 
was reported for 201 SAEs, with 14 (7%) of reported SAEs occurring on booster vaccination. 
The most serious and potentially life-threatening AE related to this vaccine are neurotropic 
(YEL-AND) and viscerotropic disease (YEL-AVD). CDC reports YEL-AND observational data74 
for 462 million vaccine doses distributed with 218 YEL-AND cases: vaccination type was known 
for 110 of the YEL-AND cases, of which 3 (3%) were booster vaccinations. CDC also reports 
YEL-AVD observational data for 437 million vaccine doses distributed with 72 YEL-AVD 
cases;74 vaccination type was known for 31 of the YEL-AVD cases, of which 1 (3%) was a 
booster vaccination. AEs are least frequently reported in 20-49 year-old vaccinees. 
 
14.3.2 Blood Draw Risks 
 
Risks associated with drawing blood from the arm include some pain when the needle is 
inserted and a small risk of bruising and/or infection at the place where the needle enters the 
arm.  Some people may experience lightheadedness, nausea, and/or fainting. 
 
14.4 
Study Benefits 
 
The proposed studies are not expected to be of direct benefit to study subjects. However, their 
participation in these studies may further understanding of the mechanisms underlying immunity 
to yellow fever virus. 
 
14.5 
Confidentiality 
 
Demographic data (age, sex) arbovirus exposure history will be obtained by the study 
coordinator using screening and intake questionnaires developed by the PI and study 
coordinator. The study coordinator and PI will have access to individually identified private 
information about human subjects. All subject data collected as part of this study are subject the 
Health Information Portability and Accountability Act (HIPAA).  
 
14.6 
Disclosure of Data 
 
The investigator, his or her staff and associates, and the appropriate regulatory agencies may 
use the information included in this protocol as necessary for the conduct of the study and the 
safety of subjects. 
 
14.7 
Publication of Research Findings 
 
Manuscript(s) and abstract(s) prepared from the data collected during this trial will be prepared 
by the study investigators. Results of the study trial will be published without restriction. 
 
15 
SUBJECT COMPENSATION 
 
Subjects will be compensated $50 for each completed visit including the screening visit. 
 
The total subject stipend for completing the study will be $500.   
 
 
16 
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