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Safety, immunogenicity, and lot-to-lot consistency of a quadrivalent inactivated influenza vaccine in children, adolescents, and adults: A randomized, controlled, phase III trial
Josefina B. Cadorna-Carlosa1, Terry Nolan13'*'1, Charissa Fay Borja-Taborac, Jaime Santosd, M. Cecilia Montalbane, Ferdinandus J. de Loozef g, Peter Eizenbergh, Stephen Hall1, Martin Dupuyj, Yanee Hutagalungk, Stéphanie Pépinj, Melanie Savillej
a University of the East Ramon Magsaysay Memorial Medical Center, 64 Aurora Boulevard, Barangay Dona Imelda, Quezon City, Philippines b Melbourne School of Population and Global Health, University of Melbourne, 207 Bouverie Street, Melbourne, Victoria 3010, Australia c Research Institute for Tropical Medicine, Corporate Ave, Muntinlupa City, Metro Manila, Philippines d Philippine Children's Medical Center, Agham Road, Quezon City, Metro Manila, Philippines e Our Lady of Remedies Clinic, Manila, Philippines f AusTrials, 14 Primrose Street, Sherwood, Queensland 4075, Australia g School of Medicine, University of Queensland, Queensland 4072, Australia h Doctors of Ivanhoe, 131 Upper Heidelberg Rd, Ivanhoe, Victoria 3079, Australia i Emeritus Research, 291 Wattletree Rd, Malvern East, Victoria 3145, Australia j Sanofi Pasteur, 1541 Avenue Marcel Mérieux, 69280 Marcy-l'Étoile, France k Sanofi Pasteur, 6 Raffles Quay, 048580, Singapore
ABSTRACT
Background: Inactivated quadrivalent influenza vaccine (IIV4) containing two influenza A strains and one strain from each B lineage (Yamagata and Victoria) may offer broader protection against seasonal influenza than inactivated trivalent influenza vaccine (IIV3), containing a single B strain. This study examined the safety, immunogenicity, and lot consistency of an IIV4 candidate.
Methods: This phase III, randomized, controlled, multicenter trial in children/adolescents (9 through 17 years) and adults (18 through 60 years) was conducted in Australia and in the Philippines in 2012. The study was double-blind for IIV4 lots and open-label for IIV4 vs IIV3. Children/adolescents were randomized 2:2:2:1 and adults 10:10:10:1 to receive one of three lots of IIV4 or licensed IIV3. Safety data were collected for up to 6 months post-vaccination. Hemagglutination inhibition and seroneutralization antibody titers were assessed pre-vaccination and 21 days post-vaccination.
Results: 1648 adults and 329 children/adolescents received IIV4, and 56 adults and 55 children/adolescents received IIV3. Solicited reactions, unsolicited adverse events, and serious adverse events were similar for IIV3 and IIV4 recipients in both age groups. Injection-site pain, headache, malaise, and myalgia were the most frequently reported solicited reactions, most of which were mild and resolved within 3 days. No vaccine-related serious adverse events or deaths were reported. Post-vaccination antibody responses, seroconversion rates, and seroprotection rates for the 3 strains common to both vaccines were comparable for IIV3 and IIV4 in both age groups. Antibody responses to IIV4 were equivalent among vaccine lots and comparable between age groups for each of the 4 strains. IIV4 met all European Medicines Agency immunogenicity criteria for adults for all 4 strains.
Conclusions: In both age groups, IIV4 was well tolerated and caused no safety concerns, induced robust antibody responses to all 4 influenza strains, and met all EMA immunogenicity criteria for adults. Clinical trial registry number: NCT01481454.
© 2015 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND
license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
* Corresponding author. Tel.: +61 3 8344 9350. E-mail addresses: jccarlosmd@yahoo.com (J.B. Cadorna-Carlos), t.nolan@unimelb.edu.au (T. Nolan), cbtabora@yahoo.com (C.F. Borja-Tabora), jimson2@me.com (I.Santos), csmontalban@yahoo.com (M.C. Montalban), freddelooze@austrials.com.au (F.J. de Looze), peter@dr131.com.au (P. Eizenberg), stephenhall@emeritusresearch.com (S. Hall), Martin.Dupuy@sanofipasteur.com (M. Dupuy), buddy.bello@gmail.com (Y. Hutagalung), Stephanie.Pepin@sanofipasteur.com (S. Pépin), Melanie.Saville@sanofipasteur.com (M. Saville).
1 These authors contributed equally to this work. http://dx.doi.org/10.1016/j.vaccine.2015.03.065
0264-410X/© 2015 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
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ARTICLE INFO
Article history:
Received 24 September 2014 Received in revised form 22 March 2015 Accepted 23 March 2015 Available online 2 April 2015
Keywords:
1nfluenza
Vaccine
Quadrivalent
Trivalent
1mmunogenicity
Safety
1. Introduction
Inactivated quadrivalent influenza vaccines (IIV4s) contain inactivated split virions from two influenza A virus strains (H1N1 and H3N2) and from the two B lineage strains (Yamagata and Victoria) [1-3]. Each year, the exact strains to be included in the quadrivalent and trivalent seasonal influenza vaccines are determined by the World Health Organization based on influenza disease surveillance data from the previous year [4]. Because the two influenza B lineages have co-circulated globally for over a decade, persons vaccinated with trivalent inactivated influenza vaccine (IIV3), which contains only one B strain, have been left unprotected against the B lineage not included in the vaccine [1,2]. IIV4 offers improved coverage over IIV3 and is expected to substantially reduce illness and hospitalizations due to influenza B [5,6]. Several quadrivalent influenza vaccines have been recently licensed for use in the US and are either approved or awaiting approval in Europe [3,4]. These vaccines have been consistently shown to be as safe and as immunogenic as their trivalent counterparts [7-12].
A recent phase III trial conducted in France during the 2011/2012 influenza season demonstrated the safety and immunogenicity of an IIV4 candidate in healthy adults [12]. Here, we report the results of a phase III trial investigating the safety, immunogenicity, and lotto-lot consistency of this IIV4 candidate in children, adolescents, and adults, and the first results for this vaccine in children and adolescents.
2. Materials and methods
2.1. Study design
This was a phase III, randomized, controlled, multicenter trial in children/adolescents (9 through 17 years of age) and adults (18 through 60 years of age) performed at six centers in Australia and four centers in the Philippines in 2012 (ClinicalTrials.gov identifier: NCT01481454). The study was double-blind for IIV4 lots and open-label for IIV4 vs IIV3. The primary objective was to describe the safety profiles (injection-site reactions and systemic events) of IIV4 and IIV3 during the 21 days following vaccination and the serious adverse events (SAEs) for 6 months in all adult and child/adolescent participants. Secondary objectives were to demonstrate compliance of IIV4 with European Medicines Agency (EMA) immunogenicity criteria in adults, demonstrate the immunogenic consistency of three lots of IIV4, and describe postvaccination seroneutralization titers in children/adolescents. The study was approved by the independent ethics committee and/or institutional review board responsible for each study site and was carried out in accordance with International Conference on Harmonization Guidelines for Good Clinical Practice, the Declaration of Helsinki, and with all national and local ethical requirements. Written informed consent was obtained from all adult participants and from all parents or legal representatives of 9-17-year-old participants included in the trial. Some 9-17-year-old participants also provided written informed assent where required by local regulations.
2.2. Study population
Individuals 9-60 years old who had not been previously vaccinated against influenza with a 2012 Southern Hemisphere formulation or a 2011-2012 Northern Hemisphere formulation in the previous 6 months were considered for study inclusion. They were excluded if they had received another vaccination within 4 weeks before inclusion; were allergic to or had a history of a serious adverse reaction (AR) to any influenza vaccine; had a known or
suspected congenital or acquired immunodeficiency; had moderate or severe acute illness/infection or a temperature > 38.0 °C; or had recently received immunosuppressive or corticosteroid treatment, immune globulins, blood, or blood-derived products. Women were excluded if they were pregnant, lactating, or of childbear-ing potential and not using adequate birth control. Enrollment was stratified by age at each site into children/adolescents 9 through 17 years old and adults 18 through 60 years old.
2.3. Randomization
Adult participants were randomized 10:10:10:1 and children/adolescents 2:2:2:1 to be immunized with one of three lots of 11V4 or with licensed 11V3. Participants were randomized using the permuted block method with stratification by site and age group and were assigned to the treatment groups via an interactive voice or web response system. All vaccines were administered by intramuscular injection with a 16-mm, 25-gauge needle. The study was open-label for receipt of either 11V4 or licensed 11V3 and was doubleblind for 11V4 lots so that neither the investigator nor the subject or subject's legal representative knew which lot was administered. Blood samples were collected before vaccination (day 0) and 21 days after vaccination.
2.4. Vaccines
All vaccines were inactivated, split-virion preparations containing 15 |ig hemagglutinin per strain in a total volume of 0.5 mL. 11V4 lot 1 (batch S4361), lot 2 (batch S4362), and lot 3 (batch S4363) contained the A/California/07/2009 (H1N1), A/Victoria/210/2009 (H3N2), and B/Brisbane/60/2008 (Victoria lineage) strains (which were among those recommended for the 2011/2012 Northern Hemisphere and the 2012 Southern Hemisphere formulations), and the B/Florida/04/2006 strain (Yamagata lineage). The licensed 11V3 (batch H0290) was the 2011/2012 Northern Hemisphere formulation ofVaxigrip® (Sanofi Pasteur, Marcy-l'Étoile, France), which contained the A/California/07/2009 (H1N1), A/Victoria/210/2009 (H3N2), and B/Brisbane/60/2008 (Victoria lineage) strains.
2.5. Hemagglutination inhibition (HAI) assay
The HA1 assay was performed as previously described [11]. The highest serum dilution resulting in complete inhibition of hemag-glutination was determined in duplicate for each sample. The titer for each strain was calculated as the geometric mean of the reciprocal of the duplicate values obtained for each sample. The lower limit of quantitation was a titer of 10, which is the reciprocal of the lowest dilution used in the assay. Samples with HA1 antibody titers below 10 were assigned a titer of 5. The seroprotection rate for each group was the percentage of participants with a titer >40. The sero-conversion rate for each group was the percentage of participants with either a pre-vaccination titer <10 and a post-vaccination titer >40 or a pre-vaccination titer >10 and a >4-fold increase in titer at day 21.
2.6. Seroneutralization assay
The seroneutralization assay was a microneutralization assay based on methods previously described by the World Health Organization and the US Centers for Disease Control and Prevention [13,14]. Serially diluted, heat-inactivated serum samples from vaccinated child/adolescent participants were pre-incubated with a fixed amount of influenza A or B virus prior to adding the serum-virus mixture to Madin-Darby canine kidney cell cultures. After overnight incubation, viral nucleoprotein production in the infected cells was measured by enzyme-linked immunosorbent
assay using a monoclonal antibody specific to the nucleoprotein of either influenza A or B. Reduced or absent infectivity indicated the presence of influenza virus-specific neutralizing antibodies in human sera. The neutralization titer expressed by the reciprocal (1/dilution) was calculated by the intersection of the optical density curve of the test sample and the 50% neutralization point of the control optical density curve. The lower limit of quantitation was a titer of 10, which was the reciprocal of the lowest dilution used in the assay. Samples with neutralizing antibody titers below 10 were assigned a titer of 5.
2.7. Reactogenicity and safety
Solicited injection-site reactions (pain, erythema, swelling, induration, ecchymosis) and systemic reactions (fever, headache, malaise, myalgia, shivering) were recorded for 7 days after each vaccination. Unsolicited adverse events (AEs) and SAEs were collected according to the International Committee for Harmonization Guideline for Clinical Safety Data Management: Definitions and Standards for Expedited Reporting. Unsolicited AEs were collected for 21 days after vaccination. Immediate unsolicited adverse reactions (ARs) were those occurring within 30 min of vaccination and were considered to be related to the vaccination. SAEs and AEs of special interest were collected for 6 months after vaccination. AEs of special interest included anaphylaxis, Guillain-Barré syndrome, encephalitis, myelitis, neuritis, convulsions, and vasculitis. AEs and SAEs were classified by the center investigators as related or unrelated to the study vaccines.
2.8. Sample size determination
A total of 1980 participants (330 children/adolescents and 1705 adults) were planned to receive 11V4. This was estimated to allow AEs with a true incidence of 0.15% overall or 0.90% in children/adolescents to be detected with a probability of 95%. Based on simulations, 660 participants in each 11V4 lot group would provide 90% power to demonstrate lot-to-lot consistency for immunogenicity with an alpha of 5%.
2.9. Statistical analysis
Statistical analysis was performed using SAS version 9.2 (SAS Institute, Cary, NC). Safety was assessed in all participants who received a study vaccine according to the vaccine received. Lot-to-lot consistency was assessed in all participants who completed the study according to protocol. Lot-to-lot equivalence for each strain was demonstrated if the age group-stratified two-sided 95% confidence interval (CI) of the post-vaccination HA1 geometric mean antibody titer (GMT) ratio for that strain in the two lots being compared was between 0.67 and 1.5. Age group-stratified 95% C1s were calculated using an analysis of variance model (type 11 analysis) of log10-transformed titers. The 95% C1s for GMTs and GMT ratios (HA1 and seroneutralization) were calculated from the Student's t distribution of log10-transformed titers.
3. Results
3.1. Study participants
Between March 19, 2012 and April 28, 2012,1390 participants in the Philippines and 700 participants in Australia were enrolled and randomized (Fig. 1). The last follow-up visit was on October 31,2012. Of the 2090 participants, 1977 were vaccinated with 11V4 (1648 adults and 329 children/adolescents) and 111 with 11V3 (56 adults and 55 children/adolescents). A total of 2071 participants (1962 11V4, 109 11V3) completed the study. Sex ratios were nearly
equivalent in the adult 11V3 group and in both child/adolescent groups, however the adult 11V4 group contained more women (61.4%) than men (Supplementary Table 1). Within each age group, mean ages and influenza vaccination histories were similar for the two vaccine groups. Most of the participants in each group were Asian.
3.2. Reactogenicity and safety
Solicited reaction frequencies were similar for 11V4 and 11V3 in adults (11V4: 61.2%, 95% C1 [58.8; 63.6]; 11V3: 57.1%, 95% C1 [43.2; 70.3]) as well as in children/adolescents (11V4: 66.6%, 95% C1 [61.2; 71.6]; 11V3: 67.3%, 95% C1 [53.3; 79.3]) (Fig. 2). Reactogenicity profiles were also similar for the three lots of 11V4 in both age groups (data not shown). 1n all groups, pain was the most common solicited injection-site reaction, whereas headache, malaise, and myalgia were the most common solicited systemic reactions. Almost all injection-site (99.1%) and systemic reactions (96.2%) were mild or moderate in severity and most (88.2%) resolved within 3 days. Although 19 (1.2%) adult 11V4 recipients reported grade 3 headache and 16 (1.0%) reported grade 3 malaise, all other types of grade 3 reactions were reported by less than 1% of the 11V4 recipients in either age group. Less than 2% the 11V3 recipients reported a grade 3 injection-site reaction and none reported a grade 3 systemic reaction.
Frequencies of unsolicited AEs were also similar for both vaccines in adults (11V4: 18.0%, 95% C1 [16.1; 19.9]; 11V3: 12.5%, 95% C1 [5.2; 24.1]) and in children/adolescents (11V4: 17.6%, 95% C1 [13.7; 22.2]; 11V3: 29.1%, 95% C1 [17.6; 42.9]). Unsolicited AEs related to vaccination in both age groups were reported for less than 4% of 11V4 recipients (59/1648 adults, 6/329 children/adolescents) and less than 10% of 11V3 recipients (7/56 adults, 5/55 children adolescents). An unsolicited immediate AR (within 30 min) was reported by one adult in the 11V4 group (urticaria) and one adult in the 11V3 group (cough). Both resolved within 2 days and both participants completed the study. The most frequently reported unsolicited AE related to vaccination was injection-site pruritus, which occurred in 13 adults (0.8%) and one child/adolescent (0.3%) vaccinated with 11V4 and in one child/adolescent (1.8%) vaccinated with 11V3. Grade 3 vaccine-related ARs were reported for 4 adults in the 11V4 group (0.2%): 3 had tremors, dizziness, or cough that resolved in 2 days or less and 1 had fatigue that resolved within 4 days. A total of 13 SAEs were reported during the 6-month study period; all occurred in 11V4 recipients (12 adults and one child/adolescent) and none were considered to be vaccine-related. No participants discontinued the study due to an AE and no AEs of special interest or deaths were reported during the study.
3.3. Immunogenicity
Within each age group, prevaccination HA1 antibody titers were similar for participants receiving 11V3 and 11V4, and both vaccines increased HA1 antibody titers by post-vaccination day 21 (Fig. 3). For the three strains common to both vaccines (A/H1N1, A/H3N2, and B/Brisbane), within each age group, post-vaccination HA1 antibody responses induced by 11V4 were comparable to the responses induced by 11V3. 1n adults, mean GMTs for these three strains increased 7.3- to 10.0-fold with 11V4 and 6.6- and 9.3-fold with 11V3. 1n children/adolescents, GMTs increased 7.5- to 23.4-fold with 11V4 and 6.4- to 16.6-fold with 11V3. As expected for the 11V4-specific B/Florida strain, GMTs increased to a greater extent with 11V4 than with 11V3, both in adults (8.4-fold for 11V4 vs 3.2-fold for 11V3) and in children/adolescents (19.4-fold for 11V4 vs 3.8-fold for 11V3). Overall, post-vaccination GMTs for all 4 strains were at least as high in children/adolescents as in adults.
Fig. 1. Participant disposition. Adults (18-60 y) and children/adolescents (child/adols, 9-17 y) were enrolled and then randomized to receive one of three lots of IIV4 or IIV3. Two participants withdrew prior to vaccination. Following vaccination, 4 participants were withdrawn due to noncompliance (NC) or were lost to follow-up (LFU). Thirteen of the remaining participants had protocol violations and were removed from the Per Protocol Set. IIV3, trivalent inactivated influenza vaccine; IIV4, quadrivalent inactivated influenza vaccine.
IIV4 met all EMA criteria for adult vaccine recipients. In adults, post-vaccination seroprotection rates were > 99%, seroconversion rates were >59%, and post-vaccination/pre-vaccination GMT ratios were >7.3 for all four vaccine strains (Table 1). In addition, the lower limits of the 95% CIs for these values exceeded EMA criteria. Although there are no EMA criteria for children or adolescents, in these participants, post-vaccination seropro-tection rates were >98%, seroconversion rates were >61%, and post-vaccination/pre-vaccination GMT ratios were >7.5 for all four strains (Table 1).
Lot-to-lot equivalence was demonstrated for all three lots of IIV4 for all four strains (Table 2, Supplementary Fig. 1). The ratios of the overall post-vaccination GMTs for each pair of lots for each strain were between 0.82 and 1.10, and the 95% CIs for these GMT ratios were all between 0.67 and 1.5.
Seroneutralization titers for the three stains common to IIV3 and IIV4 (A/H1N1, A/H3N2, and B/Brisbane) were similar for the two children/adolescent vaccine groups (Supplementary Table 2). Like the HAI titers, seroneutralization responses for the B/Florida strain were substantially higher in the IIV4 group (26.5-fold increase) than in the IIV3 group (3.7-fold increase).
4. Discussion
Difficulties in predicting the global circulation of influenza B viruses have resulted in missed opportunities for protection. Because quadrivalent influenza vaccines induce antibody responses against both influenza B lineages, they offer broader
protection than trivalent influenza vaccines. This study found comparable reactogenicity and safety profiles for the IIV4 candidate and IIV3 in both adults and in children/adolescents. The study also showed that the immunogenicity of three lots of IIV4 was equivalent for all four strains.
Both vaccines were well tolerated by both age groups. Most solicited injection-site and systemic reactions with either vaccine were mild to moderate and most resolved within a few days. Although no grade 3 systemic reactions were reported for IIV3 (small sample size), a few were reported for IIV4. The most frequent of these was headache in adult IIV4 recipients (1.2%). All other grade 3 systemic reactions (fever, malaise, myalgia, and shivering) were reported by <1% of the IIV4 recipients in either age group. Similar findings in adults have been reported previously for IIV4 [7,11,12]. Frequencies of unsolicited AEs in the 21 days following vaccination were similar in adults and in children/adolescents vaccinated with IIV4 and few of these were considered to be treatment related.
Together with the previous phase III study [12], 328 children/adolescents, 2423 adults 18-60 years of age, and 784 older adults have now been vaccinated with the IIV4 candidate, for a total of 3525 recipients. This number of vaccinated participants should allow AEs with a true incidence of 0.085% overall to be observed with a probability of 95%. Among all IIV4 vaccinees, no participants have discontinued a study due to an AE and no vaccine-related SAEs, no AEs of special interest, and no safety issues or concerns have been reported. These two clinical trials demonstrate that the addition of a second B strain to IIV3 does not affect its safety,
Fig. 2. Solicited injection-site and systemic reactions. The percentages of participants reporting any solicited reaction and specific solicited reactions are shown for adults (A) and for children/adolescents (B). Results are for the Safety Analysis Set that included all participants who received a study vaccine. Erythema, swelling, induration, and ecchymosis in children 9-11 years old were considered grade 1 if >0 to <25 mm, grade 2 if >25 to <50 mm, and grade 3 if >50 mm; in participants 12-60 years old, they were considered grade 1 if >25 to <50 mm, grade 2 if >51 to <100 mm, and grade 3 if >100 mm. Fever was considered grade 1 if >38.0 °C to <38.4 ° C, grade 2 if >38.5 °Cto <38.9 °C, and grade 3 if >39.0 °C. Pain in children 9-11 years old was considered grade 1 if easily tolerated, grade 2 if sufficiently discomforting to interfere with normal behavior or activities, and grade 3 if incapacitating and preventing performance of usual activities. Pain in subjects 12-60 years old and all other events in all age groups were considered grade 1 for no interference with activity, grade 2 for some interference with activity, and grade 3 for significant and preventing daily activities. 11V3, trivalent inactivated influenza vaccine; 11V4, quadrivalent inactivated influenza vaccine. *The "Any" category shows the overall percentage of participants reporting any solicited injection-site or systemic reaction and the percentage reporting any grade 3 reaction. Grade 1 and 2 reactions are not shown for this category.
which agrees with reports for other quadrivalent inactivated and live-attenuated vaccines [8-11].
11V4 vaccination induced robust antibody responses for all four strains in both age groups. The antibody responses were similar
to those induced by IIV3 for the three strains in common and were higher with IIV4 for the B/Florida strain that was not in IIV3. In addition, IIV4 met all adult EMA immunogenicity criteria for seroprotection rates, GMT ratios, and seroconversion rates for
Table 1
Hemagglutination inhibition immunogenicity of 11V4.
Measure
EMA requirement
A/H1N1
A/H3N2
B/Brisbane
B/Florida
Adults (18 -60 y) Post-vaccination (day 21) seroprotection3 rate Post-vaccination (day 21) to pre-vaccination (day 0) GMT ratiob Rate of seroconversion or significant increased
Children/adolescents (9-17y) Post-vaccination (day 21) seroprotectiona rate Post-vaccination (day 21) to pre-vaccination (day 0) GMT ratiob Rate of seroconversion' or significant
>70% >2.5 >40%
99.3 (98.7, 99.6) 9.2(8.5,10.0)
63.6 (61.2, 65.9)
98.8 (96.9; 99.7) 12.0(10.2; 14.0)
77.7 (72.8; 82.1)
99.0 (98.4, 99.4)
7.3 (6.8, 7.9) 59.3 (56.8, 61.7)
100.0 (98.9; 100.0)
7.5 (6.3; 8.9) 61.8 (56.3; 67.1)
100.0 (99.8,100.0) 10.0 (9.2,10.8) 66.5 (64.2, 68.8)
99.4 (97.8; 99.9) 23.4(19.6; 27.8) 83.8 (79.3; 87.6)
100.0 (99.8,100.0)
8.4 (7.8, 9.0) 65.9 (63.5, 68.1)
99.4 (97.8; 99.9) 19.4(16.3; 23.2) 84.8 (80.4; 88.5)
Values are for the Other Analysis Set, which included randomized participants who received one dose of 11V4 and had valid pre- and post-vaccination HA1 titers. 11V4, quadrivalent inactivated influenza vaccine.
a Seroprotection is defined as a HA1 titer >40; values are the percent of participants in each group (with 95% C1s in brackets) with a HA1 titer >40.
b Values are the geometric means ofthe individual post-vaccination (day 21) HA1 titer/pre-vaccination(day 0) HA1 titer ratios (with 95% C1s in brackets) for the participants in each group.
c Seroconversion is defined as a pre-vaccination (day 0) HA1 titer <10 and post-vaccination (day 21) HA1 titer >40.
d Significant increase is defined as a pre-vaccination (day 0) HA1 titer >10 and post-vaccination (day 21) HA1 titer/pre-vaccination (day 0) HA1 titer ratio >4. Values are the percent of participants in each group (with 95% C1s in brackets) who either seroconverted or had a significant increase in HA1 titer.
Fig. 3. Geometric mean HAI titers for IIV4 and IIV3. HAI titers were measured on day 0 (pre-vaccination) and 21 days post-vaccination. Bars indicate the geometric mean titers (GMTs) in adults (A) and adolescents/children (B), and whiskers indicate the 95% CIs. Results shown are for the Full Analysis Set, which included all participants who received a study vaccine and had a valid post-vaccination titer result. IIV4, quadrivalent inactivated influenza vaccine; IIV3, trivalent inactivated influenza vaccine.
Table 2
11V4 lot-to-lot comparisons.
Strain Comparison GMT ratio (95% CI) Equivalence
A/H1N1 Lot 1 vs Lot 2 1.10(0.97 1.24) Yes
Lot 1 vs Lot 3 0.89 (o.79 1.01) Yes
Lot 2 vs Lot 3 0.82(0.72 0.92) Yes
A/H3N2 Lot 1 vs Lot 2 1.01 (0.90 1.14) Yes
Lot 1 vs Lot 3 0.91 (0.81 1.03) Yes
Lot 2 vs Lot 3 0.90 (0.80 1.02) Yes
B/Brisbane Lot 1 vs Lot 2 1.00 (0.89 1.12) Yes
Lot 1 vs Lot 3 1.00 (0.89 1.13) Yes
Lot 2 vs Lot 3 1.00 (0.89 1.13) Yes
B/Florida Lot 1 vs Lot 2 0.98 (0.88 1.10) Yes
Lot 1 vs Lot 3 0.95 (0.86 1.06) Yes
Lot 2 vs Lot 3 0.97 (0.87 1.08) Yes
Values are for the Per Protocol Set, which included randomized participants who completed the study according to protocol. The age-stratified GMTs for each strain for each lot are shown in Supplementary Fig. 1. GMT, geometric mean HAI titer. Lotto-lot equivalence for each strain was demonstrated if the two-sided 95% confidence interval (CI) of the post-vaccination ratio of the overall GMTs for that strain in the two lots being compared was between 0.67 and 1.5.
all four strains. Although there are currently no EMA criteria for children or adolescents, IIV4 responses tended to be higher in children/adolescents than in adults. These results suggest that a single dose of IIV4 provides sufficient immunity for adults and for children as young as 9 years old.
Although high HAI antibody titers generally correlate with protection for adults [15,16], the correlation is weaker in elderly adults [17,18] and in young children, the latter of which were shown to require HAI titers >110 to achieve 50% protection [19]. In addition, live-attenuated influenza vaccines are effective in young children but do not produce high HAI titers [20]. Hence, alternative sero-logical assays are under evaluation to identify other protection correlates. Seroneutralization assays measure the capacity of all antibodies in the serum to neutralize live influenza viruses and have the potential to be more sensitive than HAI assays, which measure antibodies against only one viral protein [21-24]. Our microneutralization assay is based on enzyme immunoassay of viral nucleoprotein rather than on cytopathic effect. Although this assay was originally developed for use with pandemic influenza viruses (e.g. A/H5N1 and A/H1N1) [13,14], it is now frequently used
to measure responses to other human influenza A and B viruses [7,25,26]. The assay is strain-specific, highly sensitive, and requires less time than neutralization assays based on cytopathic effect and the results have been shown to correlate with HAI [25]. IIV4 and IIV3 both increased seroneutralization antibody titers substantially in children/adolescents with patterns similar to the HAI assay for each strain. This assay should be studied as an alternative measure of immunogenicity in future clinical trials.
The results of our study are in accordance with the previous phase III trial assessing this IIV4 candidate [12] and with several other clinical trials [7-11], which have shown that quadrivalent vaccines are as immunogenic as their trivalent counterparts for the matched influenza strains and that the two vaccine formulations have comparable safety profiles. Thus, the addition of a second B strain to a licensed trivalent vaccine offers additional protection against influenza B, does not interfere with the immunogenicity of the three shared strains, and has little, if any, impact on vaccine safety or reactogenicity.
This was a large study sufficiently powered to observe infrequent AEs and to confirm lot-to-lot equivalence. Because we focused on these objectives for the adult IIV4 group, the other groups in our study were too small to permit other statistically reliable comparisons. Thus, the study did not allow assessment of non-inferiority or superiority of the IIV4 responses to IIV3 nor comparisons between age groups.
In conclusion, this study demonstrated that IIV4 can be reproducibly manufactured to yield a well-tolerated, safe, and immunogenic vaccine in persons 9-60 years of age and that it met all EMA immunogenicity criteria in adults. Vaccination with this IIV4 candidate rather than IIV3 offers a broader immune response to influenza B and might help reduce influenza-related hospitaliza-tions, costs, and deaths.
Acknowledgments
The authors thank all study participants and the parents of the children who participated. Thanks are extended to Dr. Maureen McKeirnan at the Brisbane South Central Research Centre, Carina Heights, Queensland, Australia for her participation in this study, and to research staff at each of the study sites in Australia: the Vaccine and Immunisation Research Group at the Murdoch Children's Research Institute, University of Melbourne (Research Manager Marita Kefford; research nurses, phlebotomists and vaccine logistics staff Sharon Trevorrow,Jacinta O'Keefe, Mairead Phelan,Jacinta Sonego, Janet Briggs, Marie West, Annmarie McEvoy, Clare Bro-phy, Jane Ryrie, and Judith Spotswood; study doctors Loretta Thorn, Nicole Rose, Naomi Clarke, and Rowena Christiansen; Principal Research Fellow and Associate Professor Jodie McVernon; and Maria Bisgnano of Melbourne Health for laboratory support); the National Centre for Immunisation Research and Surveillance at Children's Hospital at Westmead; the Brisbane South Clinical Research Centre in Carina Heights; AusTrials in Sherwood; Doctors of Ivanhoe in Ivanhoe; and Emeritus Research in Malvern East; and to those in the Philippines: the Lingga Health Center in Calamba Laguna City; the Research Institute for Tropical Medicine in Muntinlupa City; the Philippine Children's Medical Center in Quezon City; and the Our Lady of Remedies Clinic in Manila. The authors thank Stephen W. Hildreth, Branda T. Hu, and Katherine Fries at Sanofi Pasteur's Central Clinical Immunology Laboratory for their contributions to the serological testing. Medical writing services were provided by Kurt Liittschwager (4Clinics, Paris, France). Support for medical writing assistance and for this study was provided by Sanofi Pasteur. Sanofi Pasteur participated in study design; the collection, analysis, and interpretation of data; writing of the report; and the decision to submit the article for publication.
Contributors: JC-C and TN were coordinating investigators for the study. JC-C, TN, CBT, JS, MCM, FJdL, PE, SH, YH, and SP participated in study execution, data collection and analysis, manuscript preparation, and final approval of the manuscript. MD and MS participated in data analysis, manuscript preparation, and final approval of the manuscript. MD, SP, and MS designed the study. Conflict of interest statement: Professor Nolan's institution (Murdoch Children's Research Institute) received research grants from Sanofi Pasteur to conduct this study and has received research grants from other companies as well. Professor Nolan chaired the Australian Technical Advisory Group on Immunisation until June 2014 and he is a member of the World Health Organization Strategic Advisory Group of Experts (SAGE) on immunization. Yanee Hutagalung is a full-time employee of Sanofi Pasteur and owns stock options in Sanofi Pasteur. Martin Dupuy, Stéphanie Pépin, and Melanie Saville are full-time employees of Sanofi Pasteur. Josefina B. Cadorna-Carlos, Charissa Fay Tabora, Jaime Santos, Cecilia Mon-talban, Ferdinandus J. de Looze, Peter Eizenberg, and Stephen Hall have no conflicts of interest to declare.
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.vaccine. 2015.03.065.
References
[1] Ambrose CS, Levin MJ. The rationale for quadrivalent influenza vaccines. Hum Vaccin Immunother 2012;8(1):81-8.
[2] Belshe RB. The need for quadrivalent vaccine against seasonal influenza. Vaccine 2010;28(Suppl. 4):D45-53.
[3] Traynor K. First quadrivalent flu vaccine approved. Am J Health Syst Pharm 2012;69(7):538.
[4] Centers for Disease Control and Prevention. Prevention and control of seasonal influenza with vaccines. MMWRRecomm Rep 2013;62(RR-07):1-43.
[5] Reed C, Meltzer MI, Finelli L, Fiore A. Public health impact of including two lineages of influenza B in a quadrivalent seasonal influenza vaccine. Vaccine 2012;30(11):1993-8.
[6] Lee BY, Bartsch SM, Willig AM. The economic value of a quadrivalent versus trivalent influenza vaccine. Vaccine 2012;30(52):7443-6.
[7] Beran J, Peeters M, Dewe W, Raupachova J, Hobzova L, Devaster JM. Immuno-genicity and safety of quadrivalent versus trivalent inactivated influenza vaccine: a randomized, controlled trial in adults. BMC Infect Dis 2013;13:224.
[8] Block SL, Falloon J, Hirschfield JA, Krilov LR, Dubovsky F, Yi T, et al. Immuno-genicity and safety of a quadrivalent live attenuated influenza vaccine in children. Pediatr Infect Dis J 2012;31(7):745-51.
[9] Block SL, Yi T, Sheldon E, Dubovsky F, Falloon J. A randomized, double-blind noninferiority study ofquadrivalent live attenuated influenza vaccine in adults. Vaccine 2011;29(50):9391-7.
[10] Domachowske JB, Pankow-Culot H, Bautista M, Feng Y, Claeys C, Peeters M, et al. A randomized trial of candidate inactivated quadrivalent influenza vaccine versus trivalent influenza vaccines in children aged 3-17 years. J Infect Dis 2013;207(12):1878-87.
[11] Greenberg DP, Robertson CA, Noss MJ, Blatter MM, Biedenbender R, Decker MD. Safety and immunogenicity of a quadrivalent inactivated influenza vaccine compared to licensed trivalent inactivated influenza vaccines in adults. Vaccine 2013;31(5):770-6.
[12] Pepin S, Donazzolo Y,Jambrecina A, SalamandC, Saville M. Safety and immuno-genicity of a quadrivalent inactivated influenza vaccine in adults. Vaccine 2013;31(47):5572-8.
[13] WHO Global Influenza Surveillance Network. Manual forthe laboratory diagnosis and virological surveillance of influenza. Geneva: WHO Press; 2011.
[14] Rowe T, Abernathy RA, Hu-PrimmerJ, Thompson WW, Lu X, Lim W, et al. Detection of antibody to avian influenza A (H5N1) virus in human serum by using a combination of serologic assays. J Clin Microbiol 1999;37(4):937-43.
[15] Cox RJ. Correlates of protection to influenza virus, where do we go from here? Hum Vaccin Immunother 2013;9(2):405-8.
[16] Ohmit SE, PetrieJG, Cross RT,Johnson E, MontoAS. Influenza hemagglutination-inhibition antibody titer as a correlate of vaccine-induced protection. J Infect Dis 2011;204(12):1879-85.
[17] GovaertTM, Sprenger MJ, DinantGJ, AretzK, Masurel N, KnottnerusJA. Immune response to influenza vaccination of elderly people. A randomized double-blind placebo-controlled trial. Vaccine 1994;12(13):1185-9.
[18] McElhaney JE, Xie D, Hager WD, Barry MB, Wang Y, Kleppinger A, et al. T cell responses are better correlates ofvaccine protection in the elderly. J Immunol 2006;176(10):6333-9.
[19] Black S, Nicolay U, Vesikari T, Knuf M, Del Giudice G, Della Cioppa G, et al. Hemagglutination inhibition antibody titers as a correlate of protection for inactivated influenza vaccines in children. Pediatr Infect Dis J 2011;30(12):1081-5.
[20] DaviesJR,GrilliEA. Natural or vaccine-induced antibody as a predictor of immunity in the face of natural challenge with influenza viruses. Epidemiol Infect 1989;102(2):325-33.
[21] Hancock K, Veguilla V, Lu X, Zhong W, Butler EN, Sun H, et al. Cross-reactive antibody responses to the 2009 pandemic H1N1 influenza virus. N Engl J Med 2009;361(20):1945-52.
[22] Clark TW, Pareek M, Hoschler K, Dillon H, Nicholson KG, Groth N, et al. Trial of 2009 influenza A (H1N1) monovalent MF59-adjuvanted vaccine. N Engl J Med 2009;361(25):2424-35.
[23] Greenberg ME, Lai MH, Hartel GF, Wichems CH, Gittleson C, Bennet J, et al. Response to a monovalent 2009 influenza A (H1N1) vaccine. N Engl J Med 2009;361(25):2405-13.
[24] Treanor JJ, Campbell JD, Zangwill KM, Rowe T, Wolff M. Safety and immuno-genicity of an inactivated subvirion influenza A (H5N1) vaccine. N Engl J Med 2006;354(13):1343-51.
[25] Kim SY, Kim YK, Eun BW, Kim NH, Kang EK, Lee BS, et al. Evaluation of immuno-genicity of the 2008-2009 seasonal influenza vaccines by microneutralization test. Korean J Pediatr 2012;55(12):474-80.
[26] Talaat KR, Karron RA, Liang PH, McMahon BA, Luke CJ, Thumar B, et al. An open-label phase I trial of a live attenuated H2N2 influenza virus vaccine in healthy adults. Influenza Other Respir Viruses 2013;7(1):66-73.
