Randomized Comparison of the Immunogenicity of Recombinant and Egg-based Influenza Vaccines Among Healthcare Personnel in Israel
Phase: PHASE4
Conditions: Influenza
Sponsor: Centers for Disease Control and Prevention

 
 
       
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Israel HCP comparative immunogenicity RIV4 vs IIV4 protocol, v3 Dec 08, 2019 
 
Randomized Single-Blind Trial to Compare the Immunogenicity of 
Recombinant Hemagglutinin Influenza Vaccine with Conventional Egg-Based 
Influenza Vaccines among Healthcare Personnel in Israel 
 
Protocol Version 3, 08 Dec 2019 
NCT04523324 
 
 
 

 
 
       
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Israel HCP comparative immunogenicity RIV4 vs IIV4 protocol, v3 Dec 08, 2019 
 
TABLE OF CONTENTS 
Abbreviations ...................................................................................................................................4 
1. PROJECT SUMMARY ...............................................................................................................4 
2. STUDY SPONSOR AND INVESTIGATORS ...........................................................................5 
2.1 Study Sponsor ...................................................................................................................5 
2.2 Investigators ......................................................................................................................5 
2.3 Advisors ............................................................................................................................6 
3. INTRODUCTION .......................................................................................................................6 
3.1 Background Information ...................................................................................................6 
3.2 Justification .......................................................................................................................8 
3.3 Expected benefits from the proposed study ......................................................................9 
4. STUDY OBJECTIVES ................................................................................................................9 
4.1 Primary Objective .............................................................................................................9 
4.2 
Secondary Objectives................................................................................................10 
4.3 Exploratory Objectives ...................................................................................................10 
5. STUDY ENDPOINTS ...............................................................................................................10 
5.1 Primary Endpoints ..........................................................................................................10 
5.2 Secondary Endpoints ......................................................................................................11 
5.3 Exploratory Endpoints ....................................................................................................11 
6. METHODS ................................................................................................................................12 
6.1 Overview of Study Design ..............................................................................................12 
6.2 Study Setting and Source Population ..............................................................................13 
6.3 Participant Identification and Recruitment .....................................................................13 
6.3.1 Inclusion and Exclusion Criteria......................................................................13 
6.3.2 Pre-screening and recruitment .........................................................................14 
6.3.3 Screening, consent and enrollment questionnaire............................................15 
6.3.4 Randomization .................................................................................................15 
6.4 Study visit 1 ....................................................................................................................16 
6.4.1 Pre-vaccination Questionnaire .........................................................................16 
6.4.2 Blood Draw ......................................................................................................16 
6.4.3 Vaccine administration ....................................................................................16 
6.5 Study Visit 2 ...................................................................................................................17 
6.6 Post-vaccination safety monitoring.................................................................................17 
6.7 Data Collection ...............................................................................................................17 
7. LABORATORY PROCEDURES .............................................................................................18 
7.1 Blood Collection/Processing/Storage .............................................................................18 
7.2 Specimen Testing ............................................................................................................18 
8. VACCINE PRODUCT ..............................................................................................................19 
9. DATA ANALYSIS ....................................................................................................................20 
9.1 Power and Sample Size Considerations ..........................................................................20 
9.2 Statistical Analysis ..........................................................................................................22 

 
 
       
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10. ETHICAL CONSIDERATIONS .............................................................................................23 
10.1 Possible Risks ...............................................................................................................23 
10.2 
Remuneration ............................................................................................................24 
10.3 Provisions for protecting privacy/ confidentiality: .......................................................24 
10.4 Consent forms for study volunteers: .............................................................................24 
10.5 Protocol Deviations .......................................................................................................25 
11. SITE MONITORING PLAN ...................................................................................................25 
11.1 Study Oversight, Management, and Guidance for Decision Making ...........................25 
12. REFERENCES ........................................................................................................................26 
 
 
List of Appendices  
 
Appendix A: Recruitment 
 
Appendix B: Eligibility Screening Form 
 
Appendix C: Consent Form 
 
Appendix D: Enrollment Questionnaire 
 
Appendix E: Pre-vaccination Questionnaire/Vaccine Administration Form 
 
Appendix F: Blood Specimen Collection/Tracking Form 
 
Appendix G: Protocol Deviation Log 
 
Appendix H: Study Status Change Form 
 
Appendix I: List of High-Risk Medical Conditions 
 
Appendix J. Data Use Agreement 
 
Appendix K. Scientific Advisory Group 
Appendix L. Adverse Event Follow-Up Form 
 
 
 
 
List of Tables 
Table 1. Specimen Collection Timing, Type, Testing Purpose and Location of Testing  
Table 2. Sample size calculations to detect specified differences in SCR between RIV4 
and IIV4 at approximately 28 days post-influenza vaccination 
Table 3: List of Appendices by Relevant Study Activity 
 
 
 

 
 
       
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ABBREVIATIONS 
ADCC 
Antibody-dependent cell-mediated cytotoxicity 
GMT 
Geometric mean titers 
HCP 
Healthcare Personnel 
HA 
Hemagglutinin 
HI 
Hemagglutination Inhibition 
ILI 
Influenza-like illness 
IIV 
Inactivated Influenza Vaccine  
MDCK 
Madin-Darby Canine Kidney 
GMR 
Geometric Mean fold rise  
MN 
Microneutralization  
NA 
Neuraminidase 
NAI 
Neuraminidase antibody mediated inhibition 
rHA 
Recombinant Hemagglutinins 
RIV 
Recombinant Influenza Vaccine 
SCR 
Seroconversion rate 
SHIRI 
Study of Healthcare Personnel with Influenza and other Respiratory Viruses in Israel 
1. PROJECT SUMMARY 
Healthcare personnel (HCP) are believed to be at increased risk of influenza virus 
infection due to a higher rate of exposure compared with the general population.1 Close contact 
with patients may further result in transmission to patients.2 Vaccination is the most effective 
method of preventing influenza, but there is limited information on the effectiveness of influenza 
vaccines among HCP.3 Some studies have demonstrated reduced influenza vaccine 
immunogenicity and effectiveness among persons with a history of frequent influenza 
vaccination,4,5 emphasizing the need to examine influenza vaccine effects among HCP who may 
be frequently vaccinated or demonstrate a high baseline immune response to influenza viruses.  
The primary objective of the study is to compare humoral immune responses to a single 
dose of recombinant hemagglutinin quadrivalent influenza vaccines (RIV4) or to standard egg-

 
 
       
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based unadjuvanted quadrivalent influenza vaccines (IIV4) among HCP ≥ 18 years old who were 
vaccinated in the 2018-19 season with IIV4. The trial will be conducted at two hospital sites in 
Israel during the upcoming influenza season (2019-20) among HCP who were enrolled in the 
Study of Healthcare Personnel with Influenza and other Respiratory Viruses in Israel (SHIRI).  
The study design is a randomized, single-blind, trial. During the study, eligible HCP at 
each site who consent to participate will be randomized 1:1 to receive a single dose of RIV4 
(Flublok™ Quadrivalent by Sanofi, Inc., 45µg of HA per strain) versus a single dose of IIV4 
(VaxigripTetra™ by Sanofi, Inc., 15µg of HA per strain). Participants will be blinded to the 
vaccine that they receive; study staff will not be blinded. Eligible HCP who report already 
having been vaccinated with Vaxigrip at the time they are approached to join the study will be 
eligible to participate. All randomized participants will have blood collected just prior to 
vaccination; all participants, including those who were vaccinated with Vaxigrip outside of the 
study, will have blood collected  , approximately 28 days after to evaluate humoral immune 
responses to vaccination. Relative efficacy of single doses of study vaccines will be assessed by 
comparing immunologic responses to vaccination among participants receiving RIV4 and those 
receiving IIV4. In addition, the effect of prior vaccination in potentially modifying antibody 
immune responses to vaccine will be evaluated. 
2. STUDY SPONSOR AND INVESTIGATORS 
2.1 Study Sponsor 
Abt Associates, Washington, DC, USA 
2.2 Investigators 
Clalit Research Institute 
Ran Balicer, MD, PhD 
Avital Hirsch, MPH 
Soroka Hospital 
David Greenberg, MD 
Yaakov Dryer, MD, PhD 
Beilinson Hospital 
Alon Peretz, MD 
Consultant 

 
 
       
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Mark Katz, MD 
University of Michigan 
Emily Martin, PhD 
Adam Lauring, MD, PhD 
Josh Petrie, PhD 
Arnold Monto, MD 
U.S. Centers for Disease Control and Prevention 
Mark Thompson, PhD 
Ashley Fowlkes, MPH 
Eduardo Azziz-Baumgartner, MD 
Abt Associates 
Laura Edwards, MPH  
Angela Cheung, MPH  
 
2.3 Advisors 
Lisa Grohskopf, MD, Influenza Division, U.S. Centers for Disease Control and Prevention 
Min Levine, PhD, Influenza Division, U.S. Centers for Disease Control and Prevention 
3. INTRODUCTION 
3.1 Background Information 
 
HCP are believed to be at increased risk of influenza virus infection due to a higher rate of 
exposure compared with the general population.1 Close contact with patients may further result 
in transmission to patients.2 Vaccination is the most effective method of preventing influenza, 
and HCP were identified by the World Health Organization as a priority population to target for 
influenza vaccination.6 However, there is limited information on the effectiveness of influenza 
vaccines among HCP.3 Reports of reduced influenza vaccine effectiveness among persons with a 
history of frequent influenza vaccination in some studies4,5 and seasons7 also make it especially 
important to examine influenza vaccine effects among HCP. Recent studies of influenza vaccine 
immunogenicity among HCP have demonstrated that repeated vaccination can blunt the antibody 
response to hemagglutinin (HA)8 and neuraminidase (NA).9 

 
 
       
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Influenza vaccine efficacy varies annually depending on the match between vaccine strains 
and circulating strains, and by influenza virus type and subtype.10 Reduced influenza vaccine 
effectiveness has been observed for A/H3N2 viruses over the past several influenza seasons in 
the United States,11,12 and in Israel,13,14 and has been associated with mutations that can occur 
during the production of inactivated influenza vaccines (IIV). Specifically, circulating influenza 
viruses are adapted for growth in embryonated chicken eggs, and then grown in large scale to 
produce the vaccine.15-17 Both the adaptation process and serial passage of influenza viruses in 
chicken eggs can result in the development of mutations that have been associated with 
diminished vaccine effectiveness against both A/H3N2 and A/H1N1 viruses.17-21  
 
New influenza vaccines manufactured without the use of eggs have been licensed in the 
United States, including a recombinant HA influenza vaccine (Flublok® by Sanofi Pasteur). 
Flublok® is formulated with purified influenza HA protein from cell-culture isolates of the 
vaccine reference virus. The influenza HA is grown using baculovirus vector technology that 
efficiently expresses the recombinant HA (rHA) proteins in large volumes in an insect cell line 
from the fall armyworm. The rHA is then purified from the cells and stored in phosphate-
buffered saline (PBS). Since production of Flublok® does not use egg-adapted reference viruses, 
the rHA proteins may more closely match the vaccine reference virus for specific components in 
some seasons.22  
 
Flublok®, which contains more influenza antigen compared to most other vaccines licensed 
in the United States (45 μg vs. 15 μg of HA per strain in IIV), has been shown to be efficacious 
and immunogenic in a number of trials. In a phase III randomized placebo-controlled trial of 
trivalent Flublok® among healthy adults aged 18-49 years, Flublok® was 45% (95% CI 19-63%) 
efficacious against influenza infection during the 2007-2008 influenza season when the majority 
of circulating influenza A/H3N2 and B viruses were drifted from the vaccine strains.23 Flublok® 
was also found to be highly immunogenic for all three vaccine strains; ≥96% of participants had 
an antibody titer of >1:40 against each vaccine strain at 28 days post-vaccination.23 In a phase III 
randomized trial comparing quadrivalent Flublok® to quadrivalent Fluarix, an egg-based 
vaccine, among adults aged ≥50 years, Flublok® was 30% (95%CI 10-47%) more efficacious 
against influenza infection during the 2014-2015 influenza season, when drifted A/H3N2 viruses 
predominated.24 Flublok® was found to be well tolerated, with equivalent reactogenicity 
compared to IIV.25 

 
 
       
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Flublok® was licensed in the United States based on demonstrations of clinical efficacy and 
non-inferiority using immunogenicity outcomes according to the criteria of the Centers for 
Biologic Evaluation and Research. Sub-analyses in the clinical trials demonstrated a greater 
response to Flublok compared with IIV among previously vaccinated individuals. However, this 
same effect has not been studied among HCP, a population who are repeatedly vaccinated 
against influenza and frequently exposed to influenza viruses. It also remains unclear whether 
recombinant HA influenza vaccines that are free from mutations in the viral HA introduced 
during the egg adaptation and passage process of egg-based vaccine production are more 
effective against circulating influenza viruses compared to egg-based vaccines. Historically, the 
immunogenicity of influenza vaccines has been assessed by measuring antibody responses to 
egg-grown influenza viruses, which may be a suboptimal measure of efficacy if egg-grown 
viruses differ antigenically from circulating wild-type viruses. Studies evaluating Flublok have 
measured antibody responses to cell culture-grown viruses, but to date, there are few data 
directly comparing the immunogenicity of recombinant HA influenza vaccines to egg-based 
vaccines using the same immunogenicity outcome measures against the same antigenic targets.   
 
This randomized, single-blind trial will assess humoral responses to recombinant HA 
quadrivalent influenza vaccine (RIV4, 45μg of HA per strain) compared with conventional egg-
based quadrivalent standard dose (IIV4, 15µg of HA per strain) among HCP ≥ 18 years old using 
both cell-grown and egg-grown vaccine reference viruses as antigenic targets.    
 
The primary study hypothesis is that a single dose of RIV4 may be more immunogenic than a 
single dose of egg-based standard-dose IIV4 to vaccine virus strain components among persons 
≥ 18 years old.  
3.2 Justification 
 
Influenza vaccines are the most effective method of influenza prevention. However, the 
efficacy of licensed influenza vaccines varies annually depending on the match between vaccine 
strains and circulating strains and varies by influenza virus type and subtype. Data from the last 
several influenza seasons showing lower vaccine effectiveness against influenza A/H3N2 viruses 
combined with studies documenting challenges with growing recently circulating influenza 
A/H3N2 viruses in embryonated chicken eggs for vaccine production have raised questions 
about the impact of egg adaptation and propagation of vaccine viruses on the effectiveness of 

 
 
       
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egg-based influenza vaccines. Furthermore, studies conducted among HCP repeatedly vaccinated 
for influenza have demonstrated a reduced antibody response to vaccination with IIVs. Influenza 
vaccines, including recombinant HA influenza vaccines, made by alternative methods of 
production that avoid the egg adaptation and/or egg propagation steps are now licensed in the 
United States. Studies are needed to compare the immunogenicity of conventional egg-based 
vaccines with vaccines manufactured using alternative methods of production using standard 
assays and endpoints. Data from such studies will inform whether a preferential recommendation 
for vaccines based on a specific method of production are warranted, particularly for HCP.  
  
3.3 Expected benefits from the proposed study 
 
Influenza results in substantial morbidity and mortality, and annual vaccination remains the 
most effective method to prevent influenza and its complications. Through this study, we will 
gain a better understanding of whether RIV4 (Flublok® Quadrivalent) offers greater protection to 
HCP than the conventional egg-based IIV4 (Vaxigrip® Quadrivalent) as measured primarily by 
humoral immune responses to MDCK cell-grown vaccine strains.  
 
In the United States, both vaccines are licensed for use in adults aged >18 years based on 
phase III trials demonstrating efficacy in comparison to placebo23 or non-inferiority to 
conventional egg-based influenza vaccines. Therefore, study participants may directly benefit 
from receipt of study vaccine by receiving protection against circulating influenza viruses during 
the 2019-20 influenza season. In Israel, only IIV4 is licensed for use in adults aged >18. 
4. STUDY OBJECTIVES 
4.1 Primary Objective  
• Compare humoral immune responses to a single dose of RIV4 versus IIV4 at 
approximately 28 days after vaccination in the 2019-20 season among HCP ≥ 18 years 
old who were vaccinated in the 2018-19 season with IIV4 (defined as Group One) as 
measured by 
o hemagglutination inhibition (HI) titers for egg-grown influenza viruses A/H1N1, 
B/Yamagata, and B/Victoria, and egg-grown and cell-grown A/H3N2 
o microneutralization (MN) titers for egg-grown and cell-grown A/H3N2 viruses  

 
 
       
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4.2 Secondary Objectives  
• Compare humoral immune responses to a single dose of RIV4 versus IIV4 at 28 days 
post-vaccination in the 2019-20 season among HCP with a history of PCR-confirmed 
influenza infection during any of the three study years (2016-17 through 2018-19) 
compared to those without PCR-confirmed influenza infection (defined as Group Two);  
• Compare humoral immune responses at 28 days post-vaccination as stated in the primary 
objective but with secondary indicators of humoral immune response using MN titers for 
other virus subtypes/lineages, as appropriate;  
• Examine whether the number of prior influenza vaccinations during the preceding 10 
years, as documented by medical records, modifies humoral immune responses after a 
single dose of RIV4 or IIV4 at approximately 28 days after receipt of the 2019-20 
vaccines as measured by  
o HI titers for influenza A/H1N1, influenza B/Yamagata, and influenza B/Victoria 
strains and  
o MN titers for influenza A/H3N2 viruses 
4.3 Exploratory Objectives  
• Examine whether any association between prior vaccination and immune response to 
2019-20 vaccination is mediated by antibodies to the prior vaccine strains  
• Gather information on any adverse events following vaccination  
• Characterize humoral immune responses as measured by HI, MN titers, and other 
appropriate humoral immunity assays among subsets of enrollees of particular interest 
due to their age or demographic characteristics, health status, and/or occupational roles 
and responsibilities. 
5. STUDY ENDPOINTS  
5.1 Primary Endpoints 
 
The primary endpoints will be assessed by the following laboratory tests. The choice of 
laboratory assay may depend in part on how the 2019-2020 vaccine components and circulating 
strains affect which laboratory assay is most effective in assessing specific antigens. 

 
 
       
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• HI and/or MN responses to egg-grown vaccine reference viruses for all vaccine viruses 
for each study season at approximately 28 days, including the following endpoints 
o Seroconversion rate (SCR) defined as the proportion of participants with paired 
samples that achieved ≥4-fold rises comparing post- versus pre-vaccination titers 
o Geometric mean titers (GMT) and the ratio of post-vaccination titers between the 
two vaccines (post-vaccination GMT ratio);  
o Geometric mean-fold rise (GMR) defined as the ratio of the post-vaccination titer 
value to the pre-vaccination value  
o Elevated post-vaccination titers at thresholds greater than 1:40, 1:80, 1:160 
• MN responses to the cell-grown influenza A/H3N2 (using MDCK-SIAT or other 
appropriate cell line) vaccine reference viruses at approximately 28 days post-
vaccination, including the following endpoints: 
o SCR, post-vaccination GMT ratio, GMR, and elevated post-vaccination titers 
5.2 Secondary Endpoints 
• HI responses to egg-grown influenza A/H3N2 vaccine reference viruses at approximately 
28 days, including the following endpoints: SCR, post-vaccination GMT ratio, GMR, and 
elevated post-vaccination titers  
5.3 Exploratory Endpoints 
• HI and/or MN responses to cell-grown wild-type influenza viruses at approximately 28 
days (if appropriate), including the following endpoints  
o SCR, post-vaccination GMT ratio, GMR, and elevated post-vaccination titers 
o SCR, post-vaccination GMT ratio, and GMR as measured by MN, as appropriate 
• GMT as measured by NAI pre- and post-vaccination. 
• Indicators of immune response to vaccination based on other immunologic assays not 
listed above (as appropriate) 

 
 
       
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6. METHODS 
6.1 Overview of Study Design 
 
This is a randomized, single-blind study design. Starting in July 2019, approximately 550 to 
700 HCP from two hospitals (275-350 per hospital site) in Israel will be enrolled. Following 
completion of a written consent form, participants will complete an enrollment survey and grant 
permission for review of information collected during the SHIRI study, including vaccination 
status and immune response during the SHIRI study years. Participants will complete an 
enrollment survey and grant permission to include and integrate information collected during the 
SHIRI study into the current study’s data, including health history, influenza vaccination history, 
and immune response to vaccination during the SHIRI study years. HCP will be randomly 
assigned 1:1 to receive a single dose of IIV4 licensed in Israel (expected to be Vaxigrip® 
Quadrivalent, 15µg of HA per strain) or RIV4 (Flublok® Quadrivalent by Sanofi Pasteur, 45µg 
of HA per strain) during August-October of 2019. Participants will be blinded to the vaccine that 
they vaccine, but study staff will be aware of the vaccine. Adverse events following vaccination 
will be monitored and documented. Blood specimens will be collected prior to vaccination and 
approximately 28 days after to evaluate immune responses to vaccination.  
Eligible HCP who were already vaccinated with Vaxigrip, the vaccine that is routinely available 
in Israel, at the time they are approached to join the study, will be invited to join the study as part 
of the Vaxigrip arm. Like randomized participants, participants who were vaccinated with 
Vaxigrip outside the study will complete a written consent form, an enrollment survey and grant 
permission for review of information collected during the SHIRI study, including vaccination 
status and immune response during the SHIRI study years. As part of the consent form, 
participants will record the estimated date that they received the vaccine, and the location, and 
will authorize study staff to verify the date of current-year Vaxigrip vaccination in the Clalit 
EMR and with the hospital vaccination staff. Participants will complete an enrollment survey and 
grant permission to include and integrate information collected during the SHIRI study into the 
current study’s data, including health history, influenza vaccination history, and immune 
response to vaccination during the SHIRI study years. Blood specimens will be collected 
approximately 28 days after to evaluate immune responses to vaccination.  

 
 
       
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6.2 Study Setting and Source Population 
 
This study will be conducted among HCP who were enrolled in SHIRI for at least one year. 
SHIRI was conducted during the 2016-2017, 2017-2018, and 2018-2019 influenza seasons at the 
following two hospitals: 
Beilinson Hospital, Rabin Medical Center, Petah Tikva, Israel 
Soroka Medical Center, Beersheva, Israel 
6.3 Participant Identification and Recruitment 
 
Consent, screening, enrollment, and all study visits will take place at designated study site 
locations.  
6.3.1 Inclusion and Exclusion Criteria  
 
The following inclusion and exclusion criteria will be applied to determine eligibility for 
enrollment. 
Inclusion criteria: 
1. Aged ≥18 
2. Are currently a member of Clalit Health Services 
3. Consent to randomized receipt of influenza vaccination with either RIV4 or IIV4, or, if 
already vaccinated in the current year outside of the study, consent to receive a post-
vaccination blood draw 
4. HCP Group One participation criteria:  
o Participated in the SHIRI study and received influenza vaccination during the 
2018-19 influenza season 
5. HCP Group Two participation criteria: 
o Participated in any year of the SHIRI study and had PCR-confirmed influenza 
virus infection during any of the three study years;  
6. HCP Group One Supplemental criteria: 
o Participated in any year of the SHIRI study, not vaccinated in 2018-2019, but 
vaccinated in at least one study year; 
o Participated in the SHIRI study during all three years, regardless of vaccination 
status 
 

 
 
       
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Exclusion criteria 
1. Already received an influenza vaccine during the current influenza season 
2. Not willing or able to get the flu vaccines being used in this study;  
3. Previous hypersensitivity reaction to the study vaccines, or any vaccine, as reported by 
the subject 
4. Received any non-influenza vaccine (e.g., Hepatitis B or other vaccine recommended for 
HCP) in the 4 weeks prior to the first study visit or plans to receive a vaccine in the 4 
weeks following the first study visit 
5. Currently participating in a study that involves an experimental agent (vaccine, drug, 
biologic, device, blood product, or medication), or has received an experimental agent 
within 1 month prior to enrollment in this study, or expects to receive an experimental 
agent during participation in this study. 
6. Any condition or employment status that the potential participant or local study site 
principle investigator (PI) believes may interfere with successful completion of the study 
prior to December 2019, including being currently pregnant. 
 
Withdrawal criteria 
 
In addition to exclusion criteria identified prior to enrollment, the following criteria will be 
used to identify withdrawal from the study: 
1. Withdrawal of consent by participant for any reason 
2. As deemed necessary by the principal investigator for noncompliance or other reasons 
3. Lost to follow up defined as not returning for scheduled study visit 2 
4. Receipt of influenza vaccine outside of the study and after enrollment  
6.3.2 Pre-screening and recruitment 
 
The study will be open to HCP meeting the eligibility criteria at the two study sites. Prior to 
enrollment, study staff will identify potentially eligible HCP from SHIRI enrollment records. 
Pre-screening procedures may vary by enrollment facility. HCP will be recruited using a 
combination of methods that will be customized at each study site, including face-to-face 
meetings, phone calls, and printed material. Each site will describe the methods employed to 
recruit HCP potentially eligible for enrollment. All potential enrollees will be recorded in a 
recruitment log (Appendix A. Recruitment Form) in order to track invitations, acceptance, and 
refusal. All HCP who are interested in participation will have a screening and consent 

 
 
       
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appointment with a study physician, as required in Israel, in order to discuss the study purpose 
and procedures. 
 
From the start of enrollment, sites will make efforts to recruit HCP from the priority 
Group One. In parallel HCP from the secondary Group Two will be recruited. If Group One 
recruitment goals cannot be reached, HCP from the Group One Supplemental will be targeted. A 
minimum target of 550 participants will be enrolled, including 275 at each site (see Section 9. 
Data Analysis). The target population of HCP will include 440 HCP from Group One, and 110 
HCP from Group Two. 
6.3.3 Screening, consent and enrollment questionnaire 
 
During the screening and consent appointment, the study physician will describe the study 
briefly and then conduct a brief interview to determine study eligibility (Appendix B. Eligibility 
and Screening Form).  
 
The study physician will administer consent for eligible HCP interested in participating.  The 
consent form will explain study details, risks, and benefits, and participating HCP will be asked 
to read, sign, and date the consent form (Appendix C. Consent Form). In addition to receiving a 
copy of the full consent form, all participants will receive a brief summary of the consent form 
and an outlined schedule of study visits. All potential study participants will be free to decide if 
they want to participate in the study. The consent will include permission to access medical 
records and link data from the previous study with data collected during the current study. 
 
After consenting to the study, participants will complete a brief questionnaire about health 
status (self-perceived health status), hours and types of patient contact encountered as part of 
work duties (Appendix D. Enrollment Questionnaire).  
6.3.4 Randomization  
 
Using the list of HCP consented to participate in the study, HCP will be randomized 1:1 to 
receive one 0.5 mL intramuscular dose of Flublok® Quadrivalent or Vaxigrip Quadrivalent. 
Randomization lists will be generated by the epidemiologist and data analyst using block 
randomization to ensure equal sample size in each group.26  
 
Randomization will be conducted after enrollment is completed. Randomization will occur 
separately for participants in Group One and Group Two. 
 
Randomization will not occur for enrolled participants who were vaccinated with Vaxigrip 
outside of the study. 

 
 
       
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6.4 Study visit 1 
6.4.1 Pre-vaccination Questionnaire 
All study participants, except those who were already vaccinated outside of the study, will 
complete a second brief questionnaire to determine whether they are currently experiencing 
fever, decreased energy, muscle aches, headache, or nausea, and will be asked to rate the 
subjective severity of any symptom(s) as mild, moderate, or severe (Appendix E. Pre-
vaccination Questionnaire/Vaccine Administration Form). Appendix E will also readdress 
the study exclusion criteria. Participants reporting subjective fever (of any severity) and those 
reporting no fever but one or more symptom of moderate or severe subjective severity will not 
receive study vaccine until their illness has resolved. In this case, a follow up appointment will 
be made after resolution of the illness, at which time the participant will receive the vaccine. 
Study number assignment and pre-vaccination blood draw will take place on the day of vaccine 
administration. 
6.4.2 Blood Draw  
 
All study participants will have 20 ml of venous blood drawn for serologic testing at the 
first study visit via venipuncture in multiple serum separator tubes with clot activator and gel, 
according to CDC guidelines. The use of a butterfly needle connected to a vacutainer tube is 
recommended as the best collection method to minimize hemolysis and to reduce the risk of 
needle stick injury. As an alternative, the use of a safety needle that connects directly to the 
vacutainer tube is acceptable. Study staff will document the blood collection in Appendix F1: 
Blood Specimen Collection/Tracking Form. 
Study participants who were vaccinated with Vaxigrip outside of the study will not receive a 
blood draw during the initial study visit. For these participants, the end-of-season blood draw 
from SHIRI, which was taken at the end of the 2018-2019 influenza season, will be used as their 
“pre-vaccination” or “baseline” blood draw. 
6.4.3 Vaccine administration  
 
Participants will receive a single (0.5ml) dose of study vaccine administered in the deltoid 
muscle of the arm opposite the blood draw. Vaccine administration will be performed by a 
qualified nurse trained in the delivery of the study vaccines and documented on Appendix E. 
Pre-vaccination Questionnaire/Vaccine Administration Form. Participants will not be made 
aware of the vaccine they received until after the second blood draw (Study visit 2). 

 
 
       
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6.5 Study Visit 2  
 
The optimal timing for Study Visit 2 is 21 to 35 days after vaccination, though 21 to 62 days 
will be considered acceptable. All study participants will have 20 ml of venous blood drawn for 
serology testing. Study staff will document the blood collection in Appendix F1: Blood 
Specimen Collection/Tracking Form. 
6.6 Post-vaccination safety monitoring  
 
After receiving the vaccine, all randomized participants will be contacted by SMS or by 
telephone 2-3 days post-vaccination and asked about adverse events (Appendix I). Non-solicited 
reports of adverse events will be documented from the time of vaccination through the second 
blood draw. Adverse events reported during this time period that are deemed to be serious by a 
medical doctor will be investigated. Serious adverse event reporting forms will be completed 
according to Israel Ministry of Health guidelines. In addition, study staff will provide 
participants with a phone number to call in case they have any questions or concerns about 
possible adverse events after vaccination. 
6.7 Data Collection 
 
Linked SHIRI Data: The participants of the current study are identified from among HCP 
that previously participated in the SHIRI study during the 2016-2019 influenza seasons. Subject 
to participant consent, study investigators will link all data from the previous study with data 
collected during the current study. This reduces participation burden, since previously supplied 
information does not need to be repeated, and provides information that is essential to study 
objectives, including immune profiles in previous seasons and HI response to prior vaccinations.  
 
Enrollment Survey: Updated information on participants’ health and occupational 
responsibilities will be collected by self-report though brief surveys at enrollment. Surveys are 
designed to be self-administered electronically via the internet using the REDCap program. In 
rare cases when a participant cannot complete a survey on his or her own, study staff will 
administer the electronic survey to the participant by telephone or in-person. Data collection 
elements are outlined in the Enrollment Questionnaire (Appendix D. Enrollment 
Questionnaire). 
 
Electronic Medical Record (EMR) Extraction: For all enrolled HCP, additional data will be 
extracted from the Clalit Health Services (CHS) EMR, which contains extensive demographic 
and clinical data. Information on socio-demographic characteristics, chronic medical conditions, 

 
 
       
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medical care utilization, and influenza vaccination history (including date of receipt, vaccine 
product name, vaccine lot number, and route of administration) for the preceding 10 years will 
be extracted from CHS medical records and employee records at enrollment. 
 
7. LABORATORY PROCEDURES 
7.1 Blood Collection/Processing/Storage   
After collection, blood specimens will be transported to the local laboratory for processing 
using standard procedures. Each tube will be labeled or barcode linked with the participant ID 
number, study visit number, and date of collection. Collected blood will be stored at 4°C 
immediately either by placing the sample on ice, in a 4°C refrigerator, or in a cooler with cold 
packs. Blood may be stored at 4°C for up to 18 hours. Sometime between 1 hour and 18 hours 
after collection, the blood collection tube will be centrifuged to separate the clotted blood from 
the serum and the serum removed to a clean tube labeled or barcode linked with the study 
participant ID number, study visit number, and date of collection. The clotted blood will be 
discarded. The serum will then be divided into approximately 8-10 aliquots of 1 mL/aliquot into 
labeled tubes. After aliquoting, the serum samples will be immediately stored at -20°C or colder.  
Participant specimens will be stored indefinitely at a CDC-designated facility for future 
testing to evaluate aspects of respiratory infection and/or immune responses to influenza vaccine 
or influenza virus infection, as needed. Participants will also be asked if they would like to be 
contacted about future research studies related to this study. 
7.2 Specimen Testing 
Assessment of humoral or serologic immune response to antigens that match influenza virus 
strains that are components of the vaccines and circulating during the study period typically 
require multiple laboratory assays. We expect that primary methodologies for this effort will be 
the HI and MN assay. In recent years, MN assays have been required to assess response to cell-
grown A/H3N2 viruses, which are a better match to circulating strains and vaccine strains in 
vaccines that do not involve egg-based production, such as Flublok. Nonetheless, the 
combination of assays and antigens most relevant to the study objectives will depend on vaccine 
components, circulating strains, and performance of assays during the study period.  

 
 
       
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Additional advance serologic assays on a subgroup of specimens may be required. For 
example, to examine the extent of narrow vs. broad antibody responses and the potential over-
focusing of response on specific antibody sites, it may be necessary to quantify inhibition by 
serum of antigenic site-specific monoclonal antibodies (mAbs) to HA.17 Response to the two 
vaccine types may also differ in the extent “back boosting” of antibodies to historical vaccine 
and circulating viruses as measured by conducting an antibody landscape.27 
Sera will be tested by HI and MN to measure antibody titers against cell-grown and egg-
grown vaccine reference viruses at approximately 28 days post-vaccination compared to baseline 
samples drawn at day 0 prior to vaccination during each study season. Sera may also be tested by 
HI and MN against cell-grown wild-type influenza strains, as appropriate. MDCK-SIAT cells or 
another appropriate cell line will be used to cultivate cell-grown virus strains. A subset of sera 
from each study arm may also be tested by other immunologic assays as appropriate to measure 
activity against antigens represented in the study vaccines at day 0 and at approximately 28 days 
at each site.  
8. VACCINE PRODUCT 
Two vaccine products will be used in this trial: (1) the standard-dose quadrivalent 
inactivated influenza vaccine (expected to be Vaxigrip® Quadrivalent), and (2) Flublok® 
Quadrivalent. CDC or study sites will purchase vaccine. Both vaccines are inactivated influenza 
vaccines that are approved for use in adults aged >18 years in the United States. Vaxigrip 
Quadrivalent is licensed in Israel. Flublok is not licensed in Israel. Both vaccine products will 
contain vaccine strains representative of the four strains in the 2019-2020 northern hemisphere 
formulation.  
Vaxigrip® Quadrivalent is a split-virus/subvirion vaccine manufactured by Sanofi 
Pasteur. It is formulated from influenza virus grown on embryonated chicken eggs. Vaccine 
virus is harvested, inactivated with formaldehyde, purified by zonal centrifugation using a 
sucrose gradient, split by Triton® X-100, and then further purified to a split-virus/subvirion form. 
The resulting vaccine suspension is clear and is available for use in adults as a prefilled single-
dose 0.5mL syringe or a 5mL multi-dose vial; the vaccine is administered intramuscularly. Each 
dose is formulated to contain 15μg of HA per strain. Each 0.5mL dose of vaccine also contains 
sodium phosphate-buffered isotonic sodium chloride solution, up to 100μg of formaldehyde, and 
up to 250μg of octylphenol ethoxylate. Multi-dose vials also contain the preservatives, 

 
 
       
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Israel HCP comparative immunogenicity RIV4 vs IIV4 protocol, v3 Dec 08, 2019 
 
thimerosal and mercury. The most common reactions occurring after vaccine administration in 
adults are pain at the injection site, myalgia, headache, and malaise. The vast majority of these 
reactions are mild to moderate, and severe reactions are very rare.28 
Flublok® Quadrivalent is a recombinant vaccine manufactured by Sanofi Pasteur. The 
vaccine is formulated from purified HA protein from cell-culture isolates of the vaccine 
reference virus grown using baculovirus vector technology in an insect cell line (expresSF+
®) 
from cells of the fall armyworm. Influenza HA is removed from cells using Triton® X-100 and 
then purified by chromatography. The resulting vaccine suspension is clear and is available for 
use in adults as a prefilled single-dose 0.5mL syringe for intramuscular administration. Each 
dose is formulated to contain 45μg of HA per strain. Each 0.5mL dose of vaccine also contains 
4.4mg of sodium chloride, 0.195μg of monobasic sodium phosphate, 1.3mg of dibasic sodium 
phosphate, and 27.5μg of polysorbate 20. The vaccine may also contain up to 19mcg of 
baculovirus and Spodoptera frugiperda cell proteins, up to 10ng of baculovirus DNA and 
cellular DNA, and up to 100μg of Triton X-100.29,30 The most common systemic symptoms after 
vaccine administration are injection site tenderness or pain, headache, fatigue, muscle pain, and 
joint pain. Severe reactions are very rare. The frequency of solicited symptoms and adverse 
events were comparable or often less frequent than among IIV recipients.24,25,31    
Vaccine products will not be modified for this study. The vaccines must be stored at 2° C 
to 8° C [35.6° F to 46.4° F] until needed. Vaccine products should not be frozen and should be 
protected from natural light. Documentation of proper vaccine storage will be monitored daily 
and maintained during the duration of the trial. In the event of accidental deep-freezing or 
disruption of the cold chain, study vaccine will not be administered. 
9. DATA ANALYSIS 
9.1 Power and Sample Size Considerations 
 
To guide enrollment goals, we estimated the sample size required to do a two-sided test 
with a significance level of 0.05 and a power of 80%. The Food and Drug Administration criteria 
for demonstrating non-inferiority between IIV and a new vaccine requires comparisons of both 
the GMR and the SCR. We based expected values for the SCRs and GMRs comparing RIV3 and 
IIV3 on previously published estimates from prior vaccine immunogenicity trials. We assume 
that the SCR for IIV4 is ≥40% and ≥55% for RIV4, based on a clinical trial among adults <65 
years demonstrating the SCR for IIV4 to range from 35% to 60% and RIV4 to range from 54% 

 
 
       
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Israel HCP comparative immunogenicity RIV4 vs IIV4 protocol, v3 Dec 08, 2019 
 
to 63% against influenza A subtypes (seroconversion rates reported against influenza B viruses 
reflected trivalent vaccine formulations and were not included).23,32 We intend to have the power 
to show a 30% relative increase in SCR in the RIV arm compared to the IIV arm. Using the same 
assumptions and sample size would also provide adequate statistical power to detect a difference 
in post-vaccination GMT of >2 fold between study arms if post-vaccination GMT value is >20 
for IIV4 and ≥40 for RIV4. Table 2 provides estimated sample sizes required for varying SCR 
and differences in SCR between study arms.  
 
Based on prior experience enrolling participants at the two study sites, we will assume 
that 5% of participants will drop out before study visit 2.  
 
Table 2. Sample size calculations to detect specified differences in SCR between RIV4 and 
IIV4 at approximately 28 days post-influenza vaccination 
  
SCR at 28 days post-
vaccination in IIV arm  
Relative increase (%) in SCR in 
RIV arm compared to IIV arm 
at 28 days post-vaccination 
Total enrollment for 
both study arms 
30% 
10% 
7526 
30% 
20% 
1926 
30% 
30% 
874 
40% 
10% 
4778 
40% 
20% 
1208 
40% 
30% 
540 
50% 
10% 
3130 
50% 
20% 
776 
50% 
30% 
340 
60% 
10% 
2032 
60% 
20% 
488 
60% 
30% 
206 
 

 
 
       
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Israel HCP comparative immunogenicity RIV4 vs IIV4 protocol, v3 Dec 08, 2019 
 
9.2 Statistical Analysis  
HI and MN results will be measured by four primary parameters: SCR, post-vaccination 
GMT ratio, GMR, and post-vaccination titers equal or greater than seropositive thresholds at 
1:40, 1:80, and 1:160. SCR will be defined as the proportion of participants with paired samples 
that achieved ≥ 4 fold rise comparing post vs pre-vaccination titers and post vaccination titers ≥ 
1:40. GMR will be defined as the mean of the ratio of post-vaccination HI/MN titer and pre-
vaccination HI/MN titer for each subject. 
Primary Objective 
The primary objective of the study is compare humoral antibody responses to IIV4 and 
RIV4 among Group One HCP as measured by HI (and MN, as appropriate) at baseline (day 0), 
and approximately 28 days post-vaccination. The proportion of participants that seroconverted 
(SCR) is defined as ≥ 4-fold increase comparing post- versus pre-vaccination titers. Exact 95% 
confidence intervals will be calculated and the SCR will be compared between vaccine groups 
using χ2 or Fisher exact tests.  
Due to the skewed distribution of the resulting titer values, base-2 log transformed titers 
will be used in analyses and then the summary statistics will be back-transformed to the original 
scale for presentation of results.33 The GMT is calculated for each vaccination group as the anti- 
log2(mean[log2(titeri)]), where i represents the titer value for each participant, and 95% 
confidence intervals assume a normal distribution. To allow for calculation of HI GMTs, a titer 
of 1:5 will be assigned to those with undetectable HI antibody. 
Differences in GMT and GMR will be compared between study groups using separate 
linear regression models or a generalized linear model that accounted for repeated measures.  
Odds of achieving elevated post-vaccination titers at seropositive thresholds > 1:40, 1:80, and 
1:160 will be compared between study groups by logistic regression.  
Secondary Objectives 
 
To compare immune responses among Group Two participants with a history of PCR-
confirmed influenza infection a linear regression predicting log transformed titer will be created 
with independent variables for year of influenza infection, type of influenza virus detected, study 
arm, and potential confounders. We assume confounding will be eliminated by randomization 
and that models will not be adjusted unless differences in study arms are found during univariate 
analysis. 

 
 
       
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Israel HCP comparative immunogenicity RIV4 vs IIV4 protocol, v3 Dec 08, 2019 
 
 
To assess prior vaccination status among Group One participants (number of influenza 
vaccinations received during the preceding 10 years) as an effect modifier of differences in 
GMTs, a linear regression predicting log transformed titer will be created with independent 
variables for vaccine history, study arm, and potential confounders, and an interaction term 
between study arm and vaccine history.  
 
All primary analyses will be conducted as intention-to-treat. All tests will be 2-tailed with a 
level of significance of .05.  
 
10. ETHICAL CONSIDERATIONS 
10.1 Possible Risks 
The potential discomforts from study participation include having blood drawn, 
intramuscular injection of vaccine, and possible reactions to vaccine. Blood samples will be 
taken from all participants at each study visit. Participants may experience some minimal pain 
and/or bruising at the site of the blood draw. To prevent or lessen bruising, study staff will apply 
pressure or ask participants to apply pressure to the draw site for several minutes after the blood 
draw. There is also a slight risk of infection associated with blood draws. Study staff will swab 
the site with alcohol and use sterile equipment to make infection at the site where blood is drawn 
or where vaccination is given extremely unlikely.  
Both study vaccines are licensed by the US FDA. Vaxigrip Tetra is licensed in Israel. 
Flublok is not licensed in Israel. Occasionally, adult recipients of influenza vaccines may 
develop influenza-like reactions such as fever, body aches, headache, malaise, myalgia, and/or 
nausea. If present, these symptoms usually occur soon after vaccination and may last up to 1-2 
days post vaccination. Some participants may develop reactions at the site of vaccination 
(redness, swelling, pain, or tenderness). Analgesics such as ibuprofen or acetaminophen and rest 
will generally relieve or moderate these symptoms. Acute and potentially life-threatening allergic 
reactions are also possible, though extremely unlikely; severe reactions from influenza vaccine 
are estimated to occur in < 1 per 4 million persons vaccinated. Signs of a severe allergic reaction 
may include shortness of breath, wheezing, hives, hoarseness, difficulty swallowing, swollen 
face/ tongue/ pharynx, tachycardia, and dizziness.  

 
 
       
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10.2 
Remuneration 
Participants will receive small gifts or incentives (such as a gift card for a local restaurant) at 
study milestones: 
• At completion of enrollment (complete enrollment survey);  
• At the time of pre-vaccination blood draws and vaccination; 
• At the time of the second blood draw. 
The total value of the gifts will be in the range of 100-150 shekels. 
10.3 Provisions for protecting privacy/ confidentiality:  
All data collected in this study will be kept confidential. All written information will be stored in 
locked cabinets with limited access, and electronic information will be stored on secured servers. 
Blood samples that will be sent to collaborating institutions for processing and analysis will not 
contain any personal identification information. Collaborating institutions will destroy any stored 
data after completion of the study and data analysis. During the study, only a few members of the 
research team will have access to personal identifying information about the study participants. 
All members of the research team, including the study investigators, research nurses, research 
assistants, and research coordinators, will have access to de-identified data during the study. 
Individuals who are not a part of the research team will not have access to the study information 
or data while the study is taking place. With an official letter and a clearly stated objective, 
scientists who are not a part of the research team may receive access to de-identified data, if 
approved by the Steering Committee (described below). 
10.4 Consent forms for study volunteers:  
Written informed consent will be obtained for all potential participants prior to enrollment. In 
understandable language, trained project staff, including a medical doctor, will explain study 
procedures to potential participants and discuss the advantages and disadvantages of 
participating. Participants will be given a copy of the full consent form. Participants will be 
asked to read the full consent before agreeing to be in the study and providing their signature. 
Study coordinators will emphasize the voluntary nature of the study, the possible benefits and 
outcomes, alternatives to participation, confidentiality of participation, and the participant’s right 
to refuse and/or withdraw from the study at any time. It will be explained to participants that 

 
 
       
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discontinuation of participation or choosing not to participate will not affect their professional 
standing.  
10.5 Protocol Deviations 
Study staff will report protocol deviations using a Protocol Deviation Log (Appendix G. 
Protocol Deviation) for each participant. Sites may also complete additional protocol deviation 
forms as required by their local IRBs. Sites will report deviations to their local IRBs according to 
site-specific reporting requirements.  
 
Protocol Completion or Termination: Study staff will complete a protocol completion or 
termination form for each participant at the time the participant either completes all protocol 
procedures or at the time of termination if early termination occurs (Appendix H. Study Status 
Change Form). 
 
11. SITE MONITORING PLAN 
CDC will conduct site monitoring through teleconference calls and site visits (as needed) 
to ensure that human subject protection, study procedures, laboratory procedures, vaccine 
administration, and data collection procedures are high quality and meet guidelines, and that the 
study is conducted in accordance with the protocol and sponsor standard operating procedures. 
CDC will summarize requests to address any areas for improvement or modification that are 
identified during teleconference calls or site visits through written communication to site PIs.  
11.1 Study Oversight, Management, and Guidance for Decision Making 
 
A steering committee consisting of representatives from Clalit Research Institute, the US 
CDC, Beilinson Hospital, Soroka Medical Center and the University of Michigan School of 
Public Health will each provide high level input into this project. The steering committee will 
review and approve a data usage agreement and other guidance documents to aid in decision 
making prior to the start of the study. The steering committee will be consulted on over-arching 
project issues including final protocol decisions, adjudicating any protocol deviations that might 
occur, reviewing and confirming analysis plans, and making final decisions on analyses, 
manuscripts, and authorship as needed. Upon the completion of all study deliverables and after a 

 
 
       
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suitable moratorium, external parties may request de-identified study data from the steering 
committee as specified in U.S. Government Data Sharing guidelines.  
12. REFERENCES 
1. 
Kuster SP, Shah PS, Coleman BL, et al. Incidence of influenza in healthy adults and 
healthcare workers: a systematic review and meta-analysis. PLoS One 2011;6:e26239. 
2. 
Ahmed F, Lindley MC, Allred N, Weinbaum CM, Grohskopf L. Effect of influenza 
vaccination of healthcare personnel on morbidity and mortality among patients: systematic 
review and grading of evidence. Clin Infect Dis 2014;58:50-7. 
3. 
Wilde JA, McMillan JA, Serwint J, Butta J, O'Riordan MA, Steinhoff MC. Effectiveness of 
influenza vaccine in health care professionals: a randomized trial. JAMA 1999;281:908-13. 
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McLean HQ, Thompson MG, Sundaram ME, et al. Impact of repeated vaccination on 
vaccine effectiveness against influenza A(H3N2) and B during 8 seasons. Clin Infect Dis 
2014;59:1375-85. 
5. 
Belongia EA, Skowronski DM, McLean HQ, Chambers C, Sundaram ME, De Serres G. 
Repeated annual influenza vaccination and vaccine effectiveness: review of evidence. 
Expert Rev Vaccines 2017;16:1-14. 
6. 
Hollmeyer H, Hayden F, Mounts A, Buchholz U. Review: interventions to increase 
influenza vaccination among healthcare workers in hospitals. Influenza and other respiratory 
viruses 2013;7:604-21. 
7. 
Skowronski DM, Chambers C, De Serres G, et al. Serial Vaccination and the Antigenic 
Distance Hypothesis: Effects on Influenza Vaccine Effectiveness During A(H3N2) 
Epidemics in Canada, 2010-2011 to 2014-2015. J Infect Dis 2017;215:1059-99. 
8. 
Thompson MG, Naleway A, Fry AM, et al. Effects of Repeated Annual Inactivated 
Influenza Vaccination among Healthcare Personnel on Serum Hemagglutinin Inhibition 
Antibody Response to A/Perth/16/2009 (H3N2)-like virus during 2010-11. Vaccine 
2016;34:981-8. 
9. 
Laguio-Vila MR, Thompson MG, Reynolds S, et al. Comparison of serum hemagglutinin 
and neuraminidase inhibition antibodies after 2010-2011 trivalent inactivated influenza 
vaccination in healthcare personnel. Open Forum Infect Dis 2015;2:ofu115. 

 
 
       
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10. Belongia EA, Simpson MD, King JP, et al. Variable influenza vaccine effectiveness by 
subtype: a systematic review and meta-analysis of test-negative design studies. Lancet Infect 
Dis 2016;16:942-51. 
11. Flannery B, Chung JR, Belongia EA, et al. Interim Estimates of 2017-18 Seasonal Influenza 
Vaccine Effectiveness - United States, February 2018. MMWR Morb Mortal Wkly Rep 
2018;67:180-5. 
12. Groth N, Montomoli E, Gentile C, Manini I, Bugarini R, Podda A. Safety, tolerability and 
immunogenicity of a mammalian cell-culture-derived influenza vaccine: a sequential Phase 
I and Phase II clinical trial. Vaccine 2009;27:786-91. 
13. Mandelboim M, Glatman-Freedman A, Drori Y, et al. Ineffectiveness of the 2014-2015 
H3N2 influenza vaccine. Oncotarget 2016;7:1185-92. 
14. Stein Y, Mandelboim M, Sefty H, et al. Seasonal Influenza Vaccine Effectiveness in 
Preventing Laboratory-Confirmed Influenza in Primary Care in Israel, 2016-2017 Season: 
Insights Into Novel Age-Specific Analysis. Clin Infect Dis 2018;66:1383-91. 
15. Skowronski DM, Chambers C, Sabaiduc S, et al. Interim estimates of 2014/15 vaccine 
effectiveness against influenza A(H3N2) from Canada's Sentinel Physician Surveillance 
Network, January 2015. Euro Surveill 2015;20. 
16. Skowronski DM, Janjua NZ, De Serres G, et al. Low 2012-13 influenza vaccine 
effectiveness associated with mutation in the egg-adapted H3N2 vaccine strain not antigenic 
drift in circulating viruses. PLoS One 2014;9:e92153. 
17. Zost SJ, Parkhouse K, Gumina ME, et al. Contemporary H3N2 influenza viruses have a 
glycosylation site that alters binding of antibodies elicited by egg-adapted vaccine strains. 
Proc Natl Acad Sci U S A 2017;114:12578-83. 
18. Robertson JS. An overview of host cell selection. Dev Biol Stand 1999;98:7-11; discussion 
73-4. 
19. Xu X, Kilbourne ED, Hall HE, Cox NJ. Nonimmunoselected intrastrain genetic variation 
detected in pairs of high-yielding influenza A (H3N2) vaccine and parental viruses. J Infect 
Dis 1994;170:1432-8. 
20. Raymond DD, Stewart SM, Lee J, et al. Influenza immunization elicits antibodies specific 
for an egg-adapted vaccine strain. Nat Med 2016;22:1465-9. 

 
 
       
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21. Liu F, Tzeng WP, Horner L, et al. Influence of Immune Priming and Egg Adaptation in the 
Vaccine on Antibody Responses to Circulating A(H1N1)pdm09 Viruses After Influenza 
Vaccination in Adults. J Infect Dis 2018;218:1571-81. 
22. Dunkle LM, Izikson R. Recombinant hemagglutinin influenza vaccine provides broader 
spectrum protection. Expert Rev Vaccines 2016;15:957-66. 
23. Treanor JJ, El Sahly H, King J, et al. Protective efficacy of a trivalent recombinant 
hemagglutinin protein vaccine (Flublok(R)) against influenza in healthy adults: a 
randomized, placebo-controlled trial. Vaccine 2011;29:7733-9. 
24. Dunkle LM, Izikson R, Patriarca P, et al. Efficacy of Recombinant Influenza Vaccine in 
Adults 50 Years of Age or Older. N Engl J Med 2017;376:2427-36. 
25. Izikson R, Leffell DJ, Bock SA, et al. Randomized comparison of the safety of Flublok((R)) 
versus licensed inactivated influenza vaccine in healthy, medically stable adults >/= 50 
years of age. Vaccine 2015;33:6622-8. 
26. Kang M, Ragan BG, Park JH. Issues in outcomes research: an overview of randomization 
techniques for clinical trials. J Athl Train 2008;43:215-21. 
27. Fonville JM, Wilks SH, James SL, et al. Antibody landscapes after influenza virus infection 
or vaccination. Science 2014;346:996-1000. 
28. Sanofi Pasteur, 450/477 Fluzone® Quadrivalent Southern Hemisphere. (Accessed April 13, 
2018, at 
https://www.fda.gov/downloads/BiologicsBloodVaccines/Vaccines/ApprovedProducts/UC
M305089.pdf.) 
29. Summary Basis for Regulatory Action - Flublok Quadrivalent. 2016. (Accessed April 24, 
2019, at 
https://www.fda.gov/BiologicsBloodVaccines/Vaccines/ApprovedProducts/ucm524660.htm
.) 
30. Flublok Quadrivalent Influenza Vaccine. 2019. (Accessed April 13, 2019, at 
https://www.fda.gov/downloads/BiologicsBloodVaccines/Vaccines/ApprovedProducts/UC
M619551.pdf.) 
31. Dunkle LM, Izikson R, Patriarca PA, Goldenthal KL, Cox M, Treanor JJ. Safety and 
Immunogenicity of a Recombinant Influenza Vaccine: A Randomized Trial. Pediatrics 
2018;141. 

 
 
       
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32. Baxter R, Patriarca PA, Ensor K, Izikson R, Goldenthal KL, Cox MM. Evaluation of the 
safety, reactogenicity and immunogenicity of Flublok(R) trivalent recombinant baculovirus-
expressed hemagglutinin influenza vaccine administered intramuscularly to healthy adults 
50-64 years of age. Vaccine 2011;29:2272-8. 
33. Gaglani M, Spencer S, Ball S, et al. Antibody Response to Influenza A(H1N1)pdm09 
Among Healthcare Personnel Receiving Trivalent Inactivated Vaccine: Effect of Prior 
Monovalent Inactivated Vaccine. J Infect Dis 2014;209:1705-14. 
  
 
 

 
 
       
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Table 3: List of Appendices by Relevant Study Activity 
 
 
Appendix 
 
Screening/ 
Enrollment 
 
Visit 1 (Pre-
vaccination 
blood draw, 
vaccination) 
 
Visit 2 (2nd 
blood draw 21 
to 35 days after 
vaccination)  
Post-study 
analysis 
Appendix A. Recruitment Form 
X 
 
 
 
Appendix B: Eligibility 
Screening Form 
X 
 
 
 
Appendix C: 
Consent Form 
X 
 
 
 
Appendix D: 
Enrollment Questionnaire  
X 
 
 
 
Appendix E: 
Pre-vaccination 
Questionnaire/Vaccine 
Administration Form 
 
X 
 
 
Appendix F1: 
Blood Specimen 
Collection/Tracking Form 
 
X 
X 
 
Appendix G: Protocol 
Deviation Log 
 
(X)  
(X) 
(X) 
 
Appendix H Study Status 
Change Form 
  
(X) 
(X) 
 
Appendix I: 
List of High-Risk Medical 
Conditions 
 
 
 
(X) 
Appendix J. Data Use 
Agreement 
(X) 
(X) 
(X) 
 
Appendix K. Scientific 
Advisory Group 
(X) 
(X) 
(X) 
 
Appendix I. Adverse Event 
Follow-Up Form 
 
(X) 
 
 
Parentheses indicate that forms will be completed as needed.