Methods
This Phase II, double-blind study was conducted to assess the immunogenicity, reactogenicity and safety of QIV and TIV in children aged between 18 and 47 months who had participated in a prior study (primed cohort) or who had not participated in the prior study (unprimed cohort). The study was conducted in 2 centers in Mexico.
Children included in the study were healthy, and were ineligible for inclusion if they had: received any seasonal influenza vaccine during the current season (2009–2010), or any investigational product or vaccine within the previous 30 days; a history of hypersensitivity or allergy to any vaccine component; acute illness at the time of enrolment; a history of Guillain Barré syndrome within 6 weeks of receipt of a previous inactivated influenza vaccine; an immunosuppressive condition or had received immunoglobulins or blood products within the previous 3 months.
A parent or guardian provided written informed consent for each child. The study was undertaken in compliance with Good Clinical Practice guidelines and the Declaration of Helsinki, regulatory requirements and approved by local, regional or national Institutional Review Boards at each study site. The trial is registered at ClinicalTrials.gov (NCT00985790).
Design and Objectives
The study included 2 cohorts of children. The primed cohort were recruited from a previous study conducted in the preceding influenza season (2008–2009), which assessed a thimerosal-free TIV versus a US licensed comparator in children aged 6 to 35 months (NCT00764790); participants were eligible for enrolment in the current study if they fulfilled the inclusion criteria and had received two 0.5 mL doses of thimerosal-free TIV. The unprimed cohort was recruited from the same community as that of the primed cohort, and had not participated in the previous study, and had not received at least 2 doses of influenza vaccine in the past. Vaccination began before the start of the 2009–2010 influenza season and unprimed children received two 0.5 mL doses of QIV or TIV given 28 days apart, and primed children received one 0.5 mL dose of QIV or TIV. In the primed cohort (n = 200), immunogenicity was assessed 28 days after 1 dose of TIV or QIV (Day 28), and in the unprimed cohort (n = 400), immunogenicity was assessed in half of the population (n = 200) 28 days after the first vaccination (Day 28 sub-cohort), and 28 days after the second vaccination (Day 56 sub-cohort) in the remaining half of the population (n = 200).
The primary confirmatory objective was to assess the immunogenic non-inferiority based on geometric mean titers (GMTs) 28 days after the last dose of QIV versus TIV for the 3 vaccine strains shared by the vaccines in primed and unprimed (Day 56 sub-cohort) children pooled. A secondary confirmatory objective was to assess the immunogenic superiority based on GMTs of QIV versus TIV for the B/Victoria strain in the QIV but not in the TIV, with primed and unprimed children (Day 56 sub-cohort) pooled. Other objectives were to assess: the superiority of GMTs against the B/Yamagata strain in primed versus unprimed (Day 28 sub-cohort) children after 1 dose of QIV at Day 28 (ie, assess the effect of homologous vaccine priming by the B/Yamagata strain included in the previous season's vaccine); assess the superiority of GMTs against the B/Victoria strain in primed versus unprimed (Day 28 sub-cohort) children after 1 dose of QIV or TIV at Day 28 (ie, assess the effect of heterologous vaccine priming by the B/Yamagata strain included in the previous season's vaccine but replaced by a B/Victoria strain in the current TIV); HI antibody responses after each vaccination in each group; the reactogenicity profile of each vaccine during the 7-day postvaccination periods; unsolicited adverse events (AEs) during the 28-day post-vaccination period; serious adverse events (SAEs) during a 6-month follow-up period.
Vaccines and Randomization
The study vaccines were manufactured by GlaxoSmithKline (GSK) Biologicals (Dresden, Germany), using the Fluarix™ process. The QIV and TIV were inactivated split virion vaccines containing 15 μg hemagglutinin antigen (HA) of each vaccine strain recommended by WHO for the 2009–2010 influenza season in the Northern Hemisphere: A/Brisbane/59/2007 (H1N1), A/Uruguay/716/07 (H3N2) and B/Brisbane/60/2008 (B/Victoria-lineage). The QIV also contained 15 μg HA of B/Brisbane/3/2007 (B/Yamagata lineage). The TIV contained a total of 45 μg HA and the QIV contained 60 μg HA and both vaccines were provided as 0.5 mL doses in a prefilled syringe. The primed subjects had previously received 2 doses TIV containing 15 μg HA each of A/Brisbane/59/2007 (H1N1), A/Uruguay/716/07 (H3N2) and B/Brisbane/3/2007 (B/Yamagata lineage) as recommended for the 2008–2009 Northern Hemisphere influenza season.
Randomization of primed and unprimed cohorts to receive TIV or QIV was performed by GSK Vaccines, Wavre, Belgium using an Internet-based central randomization system. Children in the primed cohort (n = 200) were randomized 1:1 to receive 1 dose of TIV or QIV, and children from the unprimed cohort (n = 400) were randomized 1:1 to receive 2 doses of QIV or TIV. Vaccines were administered as a 0.5 mL dose in the left deltoid by blinded personnel.
Immunogenicity
Blood samples were collected for immunogenicity assessments before vaccination and 28 days after each dose. In primed subjects who received 1 dose of QIV or TIV, immunogenicity was assessed on Days 0 and 28. In unprimed subjects who received 2 doses of QIV or TIV, immunogenicity assessments were performed on Day 0 and at Day 28 in half of the group (Day 28 sub-cohort), and on Day 56 in the remaining half of the group (Day 56 sub-cohort).
Antibody titers against the vaccine strains were measured in serum samples using HI assays which were performed at GSK Vaccines' laboratory using standardized procedures as previously described. GMTs were obtained by computing the anti-log of the arithmetic mean of the log-transformed inverse titers. Children with HI antibody titers ≥1:10 were considered to be seropositive. HI antibody responses were described as seroconversion rate (SCR, defined as the percentage of children who had pre- and postvaccination titers of <1:10 and ≥1:40, respectively, or a prevaccination titer of ≥1:10 and ≥4-fold increase in postvaccination titer), seroprotection rate (defined as the percentage of children with a post-vaccination titer of ≥1:40), and the seroconversion factor (defined as the geometric mean of the ratio between post- and prevaccination reciprocal HI titers).
Reactogenicity and Safety
The parents/guardians of children used diary cards to record solicited injection-site and general AEs for 7 days following vaccination. Injection-site AEs were pain, redness and swelling, and general AEs were fever, irritability/fussiness, drowsiness and loss of appetite. Fever was defined as an oral/axillary temperature ≥37.5°C or a rectal temperature ≥38°C. Intensity of solicited symptoms was graded (0–3); Grade 1 symptoms were defined as not interfering with normal activities and Grade 3 symptoms were defined as preventing normal activities (Grade 3 redness and swelling: diameter >50 mm; Grade 3 fever: temperature >39°C).
Spontaneously reported AEs were recorded for 28 days after each vaccination, and SAEs were recorded for 6 months postvaccination. Unsolicited AEs were coded using the Medical Dictionary for Regulatory Activities. All solicited injection site symptoms were considered vaccination-related, and investigators provided causality assessments for solicited general AEs and unsolicited AEs.
Statistics
A sample size of 180 children each in the QIV and TIV groups (obtained by pooling the primed cohort and half of the unprimed cohort) was estimated to provide an overall power of 80% to evaluate the primary objective of non-inferiority of GMTs against shared vaccine strains using a 1-sided, 2-sample t-test and assuming that the upper limit of the 95% confidence interval (CI) for the GMT ratio of TIV/QIV did not exceed 2. An additional 90 unprimed children in each vaccine group was needed to evaluate the secondary objectives of homologous and heterologous priming. After adjusting for a 10% drop-out rate, the target enrolment was 300 children in each TIV and QIV vaccine group (total 600).
GMTs adjusted for baseline titer were estimated using an analysis of covariance model fitted on log10-transformed postvaccination HI titer including vaccine group as a fixed effect and baseline titer as a covariate. The primary objective of immunogenic non-inferiority of QIV versus TIV for the shared vaccine strains 28 days after the last vaccination was demonstrated if the upper limit of the 95% CI for the adjusted GMT ratio of TIV/QIV was <=2 (pooled analysis of primed cohort and unprimed Day 56 sub-cohort). The secondary objective of immunogenic superiority of QIV versus TIV for the alternate-lineage B strain 28 days after the last vaccination was demonstrated if the lower limit of the 95% CI for the adjusted GMT ratio of QIV/TIV was >1 (pooled analysis of primed cohort and unprimed Day 56 sub-cohort). The immunogenic superiority of 1 dose of QIV in primed versus unprimed (Day 28 sub-cohort) children against B/Yamagata (homologous priming) and the superiority of 1 dose of QIV or TIV in primed versus unprimed (Day 28 sub-cohort) children against B/Victoria (heterologous priming) was demonstrated if the lower limit of the 2-sided 95% CI of the adjusted GMT ratio for primed/unprimed was >1.
Post-hoc analyses were performed for both B/Yamagata and B/Victoria strains (homologous and heterologous priming) of unadjusted GMTs estimated using an analysis of variance model fitted on log10-transformed postvaccination HI titer including strain by vaccine group as a fixed effect. These analyses were performed to further evaluate the effect of priming and baseline antibody titers on post-vaccination titers.
Immunogenicity was evaluated in the per-protocol immunogenicity cohort including children who met the eligibility criteria, complied with the protocol, and for whom data were available at the evaluation time point.
The frequency of solicited and unsolicited AEs was tabulated with a 95% CI in the total vaccinated cohort including all children who received at least 1 vaccination stratified by vaccine group with the primed and unprimed cohorts pooled.