BELSHE, ROBERT B. MD; GRUBER, WILLIAM C. MD
Section Editor(s): Dagan, Ron M.D.
The large enveloped RNA viruses remain the most common causes of serious infections of childhood. Influenza A and B, parainfluenza viruses and respiratory syncytial virus are the most common causes for hospitalization of infants and young children. Significant secondary diseases including otitis media (OM) often accompany these common viral infections, and viruses may be isolated from middle ear fluid in some cases. 1–3 Despite the availability of inactivated influenza virus vaccine for children and the demonstrated efficacy of this vaccine to prevent OM, inactivated influenza vaccine is infrequently used in this age group. 4
The recent demonstration that live attenuated intranasally administered vaccine is safe and effective in children and adults represents a new opportunity to reduce the impact of influenza and to prevent its complications, including pneumonia and OM in children. 5–7 This article summarizes the recent experience in children, with special emphasis on the prevention of OM and understanding the mechanisms of action of the live attenuated vaccine.
Vaccine and placebo
Trivalent cold-adapted influenza vaccine was supplied by Aviron (Mountain View, CA), frozen in single-dose intranasal applicators described below. In Year 1 the vaccine contained 106.7 tissue culture-infective dose (TCID)50/dose, and in Year 2, 107.0 TCID50/dose of each of the three attenuated strains that matched the antigens as recommended for the trivalent inactivated influenza vaccine by the Food and Drug Administration for the 1996 to 1997 and 1997 to 1998 influenza seasons, respectively. The vaccine was stored frozen at −20°C; thawed vaccine could be stored for up to 8 h at refrigerator temperature (2–8°C) before use. The placebo consisted of egg allantoic fluid containing sucrose-phosphate-glutamate and was indistinguishable in appearance or smell from the vaccine. The spray applicator consisted of a syringe-like device that was calibrated and divided for delivery of two 0.25-ml aliquots (one per nostril) as a large particle aerosol for a total delivered volume of 0.5 ml of study vaccine or placebo.
The study population comprised 1602 healthy children ages 15 to 71 months at the time of initial vaccination in Year 1. Revaccination was offered to all available subjects from September through November, 1997. Of the original study group of 1602 children, 1358 (85%) were revaccinated with a single dose of live attenuated influenza vaccine by nasal spray. There were no statistically significant differences between the age, sex, race, day-care enrollment or household compositions of vaccine and placebo groups in Year 1 or Year 2. Informed consent was obtained from a parent or guardian. Children excluded from participation included those with a history of clinically significant hypersensitivity to eggs or children with underlying chronic illnesses for whom the inactivated influenza vaccine would be recommended.
The study was prospective, randomized, double blind, placebo-controlled and multicenter in design. The primary efficacy endpoint of the revaccination study was the first episode of culture-confirmed influenza illness in each year. Analyses included evaluation of subtype-specific efficacy, and in this case, all first cases of influenza A or influenza B were counted toward the subtype-specific efficacy. Subjects were randomized in a 2:1 ratio to receive vaccine or placebo and followed through the subsequent 2 influenza seasons. Volunteers were offered revaccination in Year 2 and were not rerandomized; subjects and staff remained blinded throughout the study. In Year 1 subjects at 8 of the 10 centers primarily received 2 doses of vaccine or placebo and 1 dose at the other 2 centers. In Year 2 subjects received 1 dose of vaccine or placebo.
Two hundred three subjects participated in an immunogenicity substudy in Year 1; 159 of these children returned in Year 2, permitting characterization of strain-specific antibody responses to the vaccine. This cohort consisted of approximately the first 21 children recruited at each site for the efficacy study, and those volunteers had blood drawn before and 4 weeks after revaccination in Year 2.
A parent or a guardian of each subject was given a digital thermometer and asked to record on a diary card daily for 10 days the temperature of the subject and the occurrence of specific symptoms including decreased activity, irritability, runny nose, sore throat, cough, headache, muscle aches, chills or vomiting. Serious adverse events occurring at any time during the study were recorded by study personnel.
Surveillance for influenza illness and case definitions
Parents were contacted by telephone weekly during the influenza seasons (November through March) to remind them to notify study personnel if the subject developed symptoms suspected to be caused by influenza; these included fever, runny nose, nasal congestion, sore throat, cough, headache, muscle aches, chills, vomiting, suspected or diagnosed OM, decreased activity, irritability, wheezing, shortness of breath or pulmonary congestion. Report of any of these symptoms or signs resulted in a culture for viruses. Study sites attempted to collect viral culture specimens from symptomatic subjects within 4 days of onset of any illness. Rhesus monkey kidney tissue culture cells were inoculated with fresh respiratory secretions within 4 h of collection or as soon as possible thereafter to cultivate influenza viruses.
The case definition of influenza was any illness detected by active surveillance (as described above) that was associated with a positive culture for wild-type influenza virus. Positive viral cultures obtained within 28 days of the annual revaccination were phenotyped to determine whether isolated viruses were wild-type influenza or were vaccine strain.
Active surveillance for symptoms and signs of influenza captured a description of the illness including whether the child was seen by the primary care provider. The provider’s diagnosis and treatment were recorded. These diagnoses included lower respiratory disease and OM with or without concomitant fever and antibiotic treatment. The case definition of lower respiratory disease was any physician-diagnosed lower respiratory disease including croup, bronchitis, pneumonia or wheezing. The case definition of febrile OM was any health care provider diagnosis of OM associated with fever (either thermometer-documented or not).
Serologic and secretory antibody studies
Sera obtained from the immunogenicity cohort were stored at −20°C and assayed for the presence of hemagglutination inhibiting antibodies to the three viruses contained in the vaccine. 8 Additionally sera were assayed at Vanderbilt University for a range of hemagglutination inhibition antibodies to a panel of H3N2 viruses, including the inactivated vaccine strain A/Nanchang/933/95, the more recent isolate A/Sydney/5/97 and other divergent H3N2 viruses: A/Thessalonika/1/95, A/Russia/13919/95 and A/Johannesburg/33/94. 9 Antigens for this comparison were obtained from Dr. Roland Lewandowski (Food and Drug Administration, Bethesda, MD). Antibody titers of ≤1:4 were considered as representing seronegative children. Nasal washes were collected on the children participating in the immunogenicity substudy at Vanderbilt University and assayed for the presence of IgA, using a modified kinetic enzyme-linked immunosorbent assay as previously described. 10
In a parallel study conducted at Vanderbilt University, 31 seronegative children age 6 to 18 months were given the recommended 2 doses of inactivated trivalent influenza vaccine [Fluzone (split); Connaught Laboratories, Swiftwater, PA]. 9 Pre- and postvaccination sera from these children were used for the assessment of antibody responses to inactivated vaccine. For this comparison we tested sera from 25 children from the present study who were initially seronegative for H3N2 and compared antibody responses stimulated by 2 doses of live attenuated intranasal influenza vaccine to the antibody responses in the cohort of children described above who received 2 doses of trivalent inactivated vaccine.
Data collection and statistical analyses
Data were monitored on site and double data entered; adverse event diagnoses were COSTART (coding symbols for thesaurus of adverse reaction terms) coded 11 by Phoenix International Life Sciences (Irvine, CA). Statistical analyses used Statistical Analysis System (SAS) Version 6.12 and StatXact 2. 12, 13 Efficacy point estimates were computed as 100 · (1 − relative risk) = 100 · (1 −PV/PP);PV and PP = observed proportion of vaccine and placebo cases, respectively. Efficacy confidence intervals used Koopman’s method for the ratio of binomials. 13 The efficacy over both seasons was based on a child’s first case of influenza using a proportional hazards model and censoring children who dropped from the study between Year 1 and Year 2. The distribution of the number of illnesses per subject was compared with a Kruskal-Wallis test. Reactogenicity P values were adjusted separately for each vaccination and symptom by Bonferonni’s method. 13 Two sided P values are reported.
No serious adverse events were associated with vaccination. Transient, minor symptoms of respiratory illness were present after Dose 1 of Year 1, when more vaccinated children, relative to placebo children, exhibited mild upper respiratory symptoms (rhinorrhea or nasal congestion (on Days 2, 3, 8 and 9 postvaccine), low grade fever (on Day 2 postvaccine) or decreased activity (on Day 2 postvaccine). 5 After revaccination no significant differences in rhinorrhea, fever or decreased activity were present. 6 Despite replication of vaccine virus in the upper airway, vaccination was not associated with OM (Table 1). When concomitant medications taken within 10 days of vaccine or placebo were compared for each dose in each year, only antipyretic use was significantly different (Year 1, Dose 1, 22% for vaccinees, 15% for placebo subjects, P < 0.05). For other doses no significant difference in antipyretic use was found. For antibiotics or antihistamines/decongestants/antitussives, no significant differences between vaccine or placebo subjects were found after any dose.
In addition to hemagglutination-inhibiting (HAI) antibody to the strain of H3N2 contained in the vaccine (influenza A/Wuhan or the antigenically equivalent A/Nanchang), the H3N2 antibodies cross-reacted with the variant influenza that emerged as the major cause of illness in 1997, influenza A/Sydney/5/97. These cross-reactive antibodies were present in 98% of vaccinated subjects compared with only 60% of placebo subjects. Furthermore the geometric mean titers (GMT) of HAI antibody to A/Sydney in vaccinated subjects were significantly higher (GMT = 68) than in placebo subjects (GMT = 12, P < 0.01). This result led us to conduct a comparison of the range of antibodies induced by the live vaccine with those in young children induced by inactivated vaccine (Fig. 1). In contrast to live vaccine, young children, 6 to 18 months of age, given 2 doses of inactivated trivalent vaccine intramuscularly developed HAI antibody to the vaccine strain (A/Nanchang) but infrequently (32%) developed HAI antibody to A/Sydney or other H3N2 variants. Live vaccine induced significantly more frequent HAI responses and higher titered responses to a range of H3N2 viruses including A/Sydney/5/97 (H3N2), Thessalonika/1/95 (H3N2), Russia/13919/95 (H3N2) and Johannesburg/33/94 (H3N2) in young children compared with the inactivated vaccine. Although significant differences in age between children in the live vaccine and inactivated vaccine groups may account for this more broad immune response, live vaccine will induce antibodies in children as young as 2 months of age. 14, 15
Live attenuated vaccine was highly efficacious (Table 2); in Year 1 the vaccine had 95% efficacy vs. H3N2 (Wuhan or Nanchang-like viruses) and 91% efficacy vs. B. In Year 2 the epidemic consisted largely of a variant not contained in the vaccine, influenza A/Sydney. In Year 2 the epidemic of A/Sydney/5/97-like viruses caused 66 of 71 cases, with the remaining cases associated with A/Wuhan/359/95-like viruses (4 cases) or influenza B (1 case). The vaccine was 100% efficacious in Year 2 against strains included in the vaccine and 86% efficacious against the variant, A/Sydney/5/97. Overall during the 2 years of study, the vaccine was 92% efficacious at preventing culture-confirmed influenza (Fig. 2).
Influenza-associated OM was significantly reduced in each year of the study. In Year 1 there was only 1 case of influenza-associated OM in the vaccine group, but there were 20 cases of OM among the placebo recipients associated with culture-positive influenza (vaccine efficacy, 98%). In Year 2 only 2 cases of OM were associated with influenza in the vaccine group, but 17 occurred in the placebo recipients (vaccine efficacy, 94%). Cases of lower respiratory disease associated with culture-positive influenza were also significantly reduced in the vaccine group; only 1 case occurred in the 2 influenza seasons in the vaccine group, but there were 11 cases in the 2 influenza seasons in the placebo recipients (vaccine efficacy, 95% against influenza culture-positive lower respiratory disease).
Several measures of vaccine effectiveness were assessed as indicators of benefit from annual vaccination (Table 3). Significant reduction in all febrile illness (regardless of result of viral cultures), reduction in febrile OM and reduction in associated antibiotic use was apparent in the vaccine groups. Similarly reduction in lost day care or lost school days and reduction in lost workdays by parents were experienced in the vaccinated children. Vaccinated children also visited health care workers significantly less often.
Live attenuated influenza vaccine provided high efficacy of 92% during 2 years against culture-confirmed influenza. This interval included a year in which the influenza strains selected for inclusion in the vaccine did not closely match the circulating predominant strain, influenza A/Sydney/5/97. Vaccine also had effectiveness in reducing febrile OM and associated antibiotic use.
The high efficacy against a variant influenza strain suggests that the live attenuated vaccine provides superior immunity compared with inactivated vaccine in years when there is a poor match between vaccine and circulating viruses. Point estimate of vaccine efficacy for the live attenuated vaccine (86%) was identical with the point estimate of protection afforded by previous natural infection with A/Wuhan/369/95-like virus (86% efficacy).
By what mechanism does the vaccine work and how does it prevent OM? Viruses may be isolated from middle ear fluid in some cases and viral infections alone or in combination with bacteria may be a cause of OM that is refractory to therapy. 1–3 Prevention of febrile OM is a result of the prevention of viral infection. This likely occurs through the induction of secretory IgA antibodies in the upper respiratory tract. Seropositive children infrequently develop serum antibodies in response to the live vaccine, and yet these children were also highly protected with the vaccine. We took the opportunity in Year 1 to examine efficacy as a function of age and found that older children were also protected from natural infection. 1 Therefore the fact that serum antibody is not boosted in seropositive individuals or in adults should not discourage use of the vaccine to prevent influenza. The development of secretory IgA is an important addition to immunity against influenza. The safety, ease of administration and high efficacy of the vaccine makes it suitable for use in children annually to prevent influenza and its complications, including OM.
Question: It is known that standard inactivated influenza vaccine prevents a substantial proportion of OM. Will the intranasal influenza vaccine further prevent OM?
Dr. Belshe: The vaccine is an important tool that will prevent OM. Preliminary results from clinical trials have shown a 18 to 33% reduction in OM cases during the influenza season. This is comparable to the reduction seen with inactivated vaccine.
Question: Are other vaccines against respiratory syncytial viruses under development?
Dr. Belshe: Phase I clinical trials in infants are under way with live attenuated respiratory syncytial virus and parainfluenza virus.
Question: What is the particle size in the vaccine aerosols and does the system deliver a uniform particle size?
Dr. Belshe: The average size of the particles is 60 μm with ∼95% of particles >10 μm in diameter.
Question: Upon administration do the vaccine particles go into the lower airway or do they stay in the nose?
Dr. Belshe: In a study using radiolabeled albumin squirted through the intranasal device, all of the material was distributed in the upper airway of adults and volunteers.
Question: Regarding attack rates in immunized children, are these truly infections in immunized children or were these vaccine failures who were infected?
Dr. Belshe: Antibody studies were performed with a subset of children but we don’t really have the serum samples to examine all the vaccinated subjects. Breakthrough infections were significantly more mild in vaccinated children in terms of fever duration, 2 days in vaccinees vs. 5 days in placebo recipients.
Question: How much of the vaccine impinges on the mucosa and how much is swallowed?
Dr. Belshe: In a dose-ranging study subjects received either drops or spray. No significant difference was noted in vaccine responses in children. In adults the spray appeared to be a better immunogen than drops. It is likely that the spray is more widely distributed on the mucosal surfaces of the upper airways.
Members of the Live Attenuated Influenza Vaccine Children’s Efficacy Trials Group include: Paul M. Mendelman, M.D., Iksung Cho, M.S., Keith Reisinger, M.D., John Treanor, M.D., Ken Zangwill, M.D., Frederick G. Hayden, M.D., David I. Bernstein, M.D., Karen Kotloff, M.D., James King, M.D., Pedro A. Piedra, M.D., Stan L. Block, M.D., Lihan Yan, MS, Janet Wittes, Ph.D., Gina Rabinovich, M.D., Mark Wolff, Ph.D. and Peter Wright, M.D.
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