Immunologic Response to Influenza Vaccination in Children and Adolescents: A RCT Trial of Influenza Vaccines
Phase: PHASE4
Conditions: Influenza, Human, Immune Response
Sponsor: Richard Zimmerman MD

Official Title:  Immunologic response to influenza vaccination 
 
ClinicalTrials.gov ID (NCT number):  NCT03614975 
 
Protocol Date:  September 20, 2018 
 
 

Scientific Background: 
The Advisory Committee on Immunization Practice (ACIP) sets civilian 
immunization policy for the United States. In adults, cell-based recombinant vaccine has 
been shown to work better immunologically against the H3 strain due to lack of egg-
based adaptations; however, it is not licensed for use in children. All of the published 
efficacy trial data on Flucelvax, the cell-based influenza vaccine approved by the FDA in 
2012, were conducted when it was based on the egg-adapted strain; it first used the 
cell-based strain in the Fall of 2017 so effectiveness data have not been published on 
this update. Egg-based vaccine has only worked reasonably well against the H3 in 
preliminary effectiveness data for younger children; it did not work well against H3 in 
adult recipients due to egg-based adaptions.  This study would be among the first head-
to-head comparison of the current versions of these two different licensed influenza 
vaccines to compare the differences in antibody response among children/teens/young 
adults. Given recent changes in the formulation of Flucelvax and the lack of published 
data on this new formulation, reasonable equipoise can be assumed among these 
vaccines. 
 
Influenza is a major public health burden, each year causing millions of illnesses 
and outpatient visits and tens of thousands of hospitalizations and deaths in the U.S. 
The primary influenza prevention method is vaccination; however, influenza vaccine 
effectiveness (VE) is variable. Host factors (e.g., age and antibody landscape), 
environment (e.g., tobacco exposure), virus and vaccine characteristics, such as the 
match between the vaccine and circulating strains, contribute to VE. This project will 
quantify pre- and post influenza immune response to different influenza vaccine types 
licensed for use (e.g., eggbased, egg-free). 
 
Study Objectives: 
Using a RCT for children 4-20 years of age, two influenza vaccines will be 
assessed for their immunogenic outcomes: Flucelvax (egg-free inactivated flu shot) and 
Fluzone (egg-based inactivated flu shot). 
 
The study objectives are to determine the serologic, proteomic and/or peripheral 
blood mononuclear cells (PBMCs)/RNA responses to: 
1) cell-based quadrivalent influenza vaccine (Flucelvax) and 2) egg-based quadrivalent 
influenza vaccine (Fluzone) in a randomized controlled trial (RCT) among racially 
diverse children 4-20 years of age.  
 
Study Design and Methods: 
We will conduct a non-blinded, randomized controlled trial of influenza vaccine 
immunogenicity in children aged 4-20 years. Influenza vaccination will occur in primary 
care settings with standard medical care use according to CDC guidelines with FDA 
approved and licensed vaccines. Bloodwork will be conducted at baseline and post-
vaccination for research purposes during the vaccination season (e.g., August thru 
December). 
 

Parents of child/teen patients will be asked to complete the appropriate screening 
questionnaire which addresses inclusion and exclusion criteria in lay language where 
possible. If they do not meet all inclusion criteria, have an exclusion criterion, or are 
unwilling to complete study activities then they will not be eligible for enrollment and will 
not be consented by the clinician. The screening questionnaire may be done in person, 
by phone, by mailed form or by electronically sent form. The questionnaire takes <10 
minutes to complete. 
 
Research coordinators will meet with parents and children/teens to review study 
requirements and provide the written informed consent form for them to read through. 
Licensed doctoral level medical providers (e.g., MDs) or licensed pharmacists (e.g., 
PharmD) will be present either in person or by electronic device (e.g., face time) to 
address any parent's/child/teen's questions regarding the study and to obtain written 
consent (parents) and assent (children/teens), including written permission from parents 
to access their child’s electronic medical record (EMR) in order to obtain data related to 
medical history, medication use, and vaccination history. 
 
Randomization to licensed flu vaccine arms: Non-blinded randomization will be 
generated by computers and done as a 1:1 allocation to each of the licensed flu vaccine 
arms. Influenza vaccine will be given by clinical staff as part of the child/teen's routine 
immunization care.  Influenza vaccine information will be collected via EMR review 
and/or documented at baseline using a research form noting the type of influenza 
vaccine received (e.g., vaccine manufacturer, valence, brand name).   
 
Blood draws will be done at UPMC or Children's clinical recruitment practices.  
All participants will have a blood draw at baseline (may be during summer before 
vaccination) and at two subsequent follow-up visits. Total blood volume drawn across 
the study visits is ~90 mL (~30 mL drawn at each study visit).  If not enough blood is 
obtained during study visit we will ask participants to return for an additional blood 
draw(s) in order to obtain the volume needed for analysis in the same study visit 
window; total volume for re-draws dependent upon tube type needed to fill (e.g, CPT 
tube for PBMCs, paxgene tube for RNA, serum tube for HI). Addendum consents for 
blood re-draws will be signed prior to drawing blood. 
 
All blood samples will be collected by trained personnel with experience drawing 
blood from children. The blood samples will be collected using Vacutainer tubes, with 
uniformly colored tops according to type of processing and appropriately labeled with a 
uniform scheme (including study ID and collection time point). To support better 
understanding of the immune response following influenza vaccination, specimens will 
be banked for future studies that might involve proteomics, immunology, 
transcriptomics, or genomics.  Collection of survey data done at UPMC clinical 
recruitment practices. Survey data will be collected from all enrolled participants at the 
baseline visit. If participant time constraints limits collection of the survey at the baseline 
visit, this information will be collected at a subsequent study visit. 
 

Cells will be prepared and frozen according to current protocols. Plasma and 
cells will be stored at -80C until all specimens are collected. They will be inventoried 
and maintained in a database in the laboratory. Upon completion of sample collection 
(pre-vacc, and postvacc) samples will be sent to the funder and other contracted 
laboratories for further analysis. Peptoid and other laboratory testing will be conducted 
per current protocols.   
 
Specific to East Liberty Family Health Center (ELFHC) clinical recruitment site: 
ELFHC clinical staff's (e.g., MA/RN) activities will be limited to 1) helping to identify 
potential subjects, drawing blood on and providing influenza vaccination to consented 
and enrolled participants. Pitt research staff along with the specified MD will conduct the 
informed consent process with parents and children/teens and/or adult (18-20 years) 
participants and Pitt research staff will administer the survey at this site. Prior to staffing 
ELFHC for study recruitment, ELFHC helping with this research project will complete 
the Community Partner Research Education and Training (CPRET) prior to engagement 
in research activities.  This training will be conducted by the Principal Investigator.   
 
All participants will have a blood draw at baseline and at two subsequent follow-
up visits.  Total blood volume drawn across the study visits is ~90 mL (~30 mL drawn at 
each study visit: baseline, 2nd study visit, 3rd study visit). If another blood draw visit 
needs to be scheduled due to lack of volume collected at the prior visit (e.g., unable to 
collect total per visit mLs due to participant dehydration) or due to poor recovery of 
PBMCs, then additional blood will be collected dependent upon the type of tube 
needed. In some circumstances (e.g., inadequate vacuum draw/clotting issue/observed 
hemolysis), up to an extra tube (e.g., up through 10mL) may be drawn at any particular 
visit.  Research staff Phlebotomists are fully trained in phlebotomy with appropriate 
clinical phlebotomy experience (e.g., experience with drawing blood on children). 
Clinical site/hospital lab Phlebotomists are fully trained and employed by UPMC. 
ELFHC clinical staff are fully trained in phlebotomy and vaccination. 
 
Specific endpoints that will result in withdrawal from the study include 
non-compliance with allotted windows for visits in study protocol. Baseline 
blood draw and 2 return blood-draw visits: day 7 (range 6-9 days) and day 
21 (range 21-35 days).  Acceptability of samples with minor deviations from 
these windows will be determined by the research team on a case-by-case 
basis. Persons who do not meet these criteria will be dropped from the 
study. Onset of a new severe bleeding disorder that precludes further  
phlebotomy. If a participant misses day 7 but can still make the day 21 visit, 
they may be retained. 
 
The PI and co-investigators will monitor the progress of the research 
study, including assessments of data quality and timeliness and participant 
recruitment, accrual and retention, subject’s confidentiality, any adverse 
event data, unanticipated problems and external factors or relevant 
information that might have an impact on the safety or ethics of the study. 
The PI and co-investigators will meet on a regular basis to discuss these 

issues. Minutes will be recorded for each of the meetings which will occur on 
a bimonthly to quarterly basis, depending on the stage of the study. At the 
time of the IRB renewal the PI will submit in writing to the IRB a summary 
report of the data and safety monitoring activities from the past year.  The 
CDC and University of Pittsburgh will hold phone calls at least quarterly to 
address the progress of the research study, including assessments of data 
quality and timeliness and participant recruitment, accrual and retention, 
subject’s confidentiality, any adverse event data, unanticipated problems 
and external factors or relevant information that might have an impact on 
the safety or ethics of the study. Minutes will be recorded for each of the 
meetings. Adverse events will be reported to the HRPO per its Policies and 
Procedures Manual. 
 
Eligibility Criteria: 
Inclusion Criteria:  
1. Males and females aged 4-20 years whose prior vaccination history is available 
(which can be determined based on the medical record or state registry for the prior 
year or participation in our previous studies of childhood influenza vaccine (previous 
participants are a high priority for enrollment); 
2. Planning to receive seasonal influenza vaccination at one of our recruiting sites 
3. >= 37 pounds 
 
Exclusion Criteria:  
1. Unable or unwilling to complete all required study activities, including informed 
consent and bloodwork 
2. Already received the influenza vaccine for the current year 
3. Have a known immunocompromising condition or on an immunosuppressing 
medication (e.g., high dose steroids) 
4. Known to be pregnant 
5. Have a history of severe allergy to eggs or to influenza vaccine or any of its 
components 
 
Statistical Considerations: 
 
As an initial step in the data analysis, descriptive statistics will be generated for 
each of the variables of interest, including their distributional properties. Through 
analyses of these distributional property assessments and relevant statistical tests, the 
necessary assumptions for the planned statistical tests will be investigated. If the 
assumptions are not met for any planned statistical test, then either the data will be 
transformed or a nonparametric alternative will be used.   
 
For the analysis of the survey data, methods appropriate for the analysis of 
counts and proportions, depending on the specific question will be used. Due to the 
distributional shape in geometric mean titers (GMT), either nonparametric tests or 
transformation of the data will be needed; log transformation will be used for the primary 
endpoint. The analysis will be conducted with transformed data. The natural log of the 

GMT will be the dependent variable to normalize the distribution. The mean and 
standard deviation of log titers will be calculated. Exact tests will be used where needed 
for small cell sizes. Regression models will also be used for predictors of immunogenic 
outcomes. 
 
Power calculations show the need to enroll 40 per vaccine arm with 74 per 
vaccine arm preferred for publication; we have noted 90 enrolled per arm to account for 
possible attrition.  A one-sided, two-sample t-test with group sample sizes of 40 and 40 
achieves 98% power to detect a ratio of 2.0 when the ratio under the null hypothesis is 
1.0. The coefficient of variation on the original scale is 1.0. The significance level (alpha) 
is 0.050.  A one-sided, two-sample t-test with group sample sizes of 74 and 74 achieves 
80% power to detect a ratio of 2.0 when the ratio under the null hypothesis is 1.0. The 
coefficient of variation on the original scale is 4.0. The significance level (alpha) is 
0.050.