Title: Seasonal influenza vaccines and hurdles to mutual protection
Author: Christopher Chiu
Affiliation: National Heart and Lung Institute, Imperial College London, London W2
1PG, United Kingdom
Correspondence: Christopher Chiu ([email protected]), National Heart and
Lung Institute, Imperial College London, London W2 1PG, UK. Tel: +44-7594 3853
Word count (abstract): 150
Word count (text): 5221
ABSTRACT
While vaccines against seasonal influenza are available, major hurdles still exist that
prevent their use having any impact on epidemic spread. Recent epidemiological
data question the appropriateness of traditional vaccination timing (prior to the winter
season) in many parts of the world. Furthermore, vaccine uptake in most countries
even in high-risk populations does not reach the 75% target recommended by the
World Health Organization. Influenza viruses continually undergo antigenic variation
and both inactivated and live attenuated influenza vaccines confer only short-lived
strain-specific immunity, so annual re-vaccination is required. Improving vaccine-
induced immunity is therefore an important goal. A vaccine that could confer durable
protection against emerging influenza strains could significantly reduce onward
transmission. Therefore, further understanding of protective immunity against
influenza (including broadly cross-protective immune mechanisms such as
hemagglutinin stem-binding antibodies and T cells) offers the hope of vaccines that
can confer the long-lived heterosubtypic immune responses required for mutual
protection.
Introduction
Vaccines are one of the major successes of modern medicine. Available cheaply and
widely used, they have been instrumental in the global control of a variety of
infectious diseases and their resulting complications [1]. This has been achieved by
inducing robust, long-lasting immunity with vaccines that recapitulate the protective
immune responses seen following natural infection with the pathogens against which
they protect. However, uniquely amongst the commonly used vaccines, those against
influenza provide minimal long-term protection and require annual re-vaccination.
Therefore it has not been possible to adequately prevent the onward transmission
that results in epidemics and pandemics. Major impediments still remain to the
generation of optimal influenza vaccine-induced immunity and these are related to
both fundamental biological features of the virus and its interaction with the host.
However, recent advances in our understanding of the immune response to influenza
infection have revived hopes for a so-called “universal” vaccine, with potential to
protect both vaccinated individuals and their contacts from newly emerging virus
strains [2].
Epidemiology and delivery of influenza vaccination
Despite the availability of vaccines, influenza is still a major cause of morbidity and
mortality worldwide. Each year, 5-15% of the world’s population will suffer an
influenza infection, with an estimated 3-5 million cases of severe disease and up to
500,000 deaths [3]. This results in expenditure of up to $167 billion per annum in the
USA alone. This enormous socioeconomic burden has made influenza control a
global priority. In temperate regions, influenza is a winter illness with epidemics
during the colder months hypothesised to be related to lower temperatures, lower
humidity, or decreased solar radiation affecting virus transmissibility [4]. In addition,
host factors such as seasonal variations in immunity and behaviour including
spending more time in contained spaces may also play a role in transmission [5].
Some of these factors may also explain the monsoon seasonality seen in sub-tropical
areas and the more year-round incidence found at tropical latitudes [6]. Only recently
has this geographically distinct epidemiology been clearly recognised and this is
beginning to have an impact on the timing of delivery of influenza vaccines. In many
regions this is still determined on a pre-“winter” seasonal schedule even when no
winter seasonality is seen. This is complicated further in countries such as India,
which, although in the northern hemisphere, spans temperate, sub-tropical and
tropical zones and still receives delivery of northern hemisphere influenza vaccines in
September or October when most cases of seasonal influenza have already occurred
[7].
Even in areas where seasonal influenza epidemics are a predictable occurrence and
concerted public health campaigns take place, vaccine uptake is generally poor [8].
In the USA, where universal vaccination is recommended, overall vaccine uptake is
approximately 44% in adults and 59% in children
(http://www.cdc.gov/flu/fluvaxview/1415season.htm). Elsewhere, influenza
vaccination is targeted towards high-risk groups such as the elderly, those with
chronic health problems or immunosuppression and pregnant women. In the UK,
vaccine coverage varies according to the group in question from approximately 44%
(in pregnant women) to 73% (in older adults)
(https://www.gov.uk/government/statistics/seasonal-flu-vaccine-uptake-in-gp-
patients-in-england-winter-season-2014-to-2015). Only a few countries globally have
therefore achieved the target set by the World Health Organization of 75% vaccine
coverage in elderly adults and those with chronic health conditions [9]. The reasons
for this are complex and multi-factorial (with strategies to address having been
extensively reviewed [10–14]) but in the absence of widespread coverage, even the
best vaccines cannot hope to reduce onward transmission, let alone influenza
vaccines which are currently suboptimal.
Influenza transmission
Influenza replicates primarily in the respiratory epithelium and is transmitted by
droplets (>10µm) and aerosols (<5µm) generated by breathing, coughing or sneezing
[5]. However, the exact mechanisms and relative contribution of different
transmission modalities amongst human populations have been difficult to clearly
establish. Thus, the role of inhalation, direct contact with droplets, indirect contact via
settled particles and fomites, and conjunctival introduction remain unclear [15,16].
For example, although influenza viruses have been found to survive on surfaces for
up to several days, the risk of transmission by indirect contact, while probably low,
has not been accurately predicted [17]. Most studies have been limited by
confounders and the extent to which transmission data from animal models can be
extrapolated to humans is unknown. It is likely that a complex interplay between the
environment, viral properties including receptor specificity and host factors is
responsible for determining the likelihood of any transmission event. However, most
environmental and virus-determined factors are difficult or impossible to alter.
Furthermore, attempting to block transmission by treating infected individuals with
antivirals or behavioural modification is futile both in terms of partial effectiveness of
these interventions and in our ability to identify infected individuals at an early stage
[18]. This is further complicated by the fact that most influenza cases are
asymptomatic while continuing to shed virus. In a large community cohort study in
England from 2006-2011, approximately 18% of unvaccinated individuals were
estimated by serological diagnosis to have been infected but around 77% of these
were asymptomatic [19]. Thus modulating the host response (and in particular the
adaptive immune response) to infection by vaccinating prior to virus exposure
continues to be the best strategy for both prevention of infection and, potentially,
transmission.
Targets of adaptive immunity
An ideal influenza vaccine would induce sterilising immunity that persisted life-long
and protected against all strains of the virus. This would protect the vaccinated
individual against influenza disease and interrupt onward transmission. However,
current vaccines are unable to consistently achieve any of these aims. Two major
factors contribute to this: the rapid and continual alteration of virus surface proteins to
evade host immunity and the inability of current vaccines to induce consistently high
levels of protection [20]. In principle, protective immunity can by conferred by
stimulation of humoral (antibody-mediated) and/or cell-mediated (T cell) immunity
[21]. These adaptive immune mechanisms are characterised by their capacity to
respond more rapidly and strongly to re-encounter with pathogens via immune
memory. However, almost all successful vaccines currently rely on antibody-
mediated correlates of protection and many questions still remain unanswered about
the role that T cells can play in vaccine-induced protection.
Haemagglutinin (HA) and neuraminidase (NA), the two major surface glycoproteins,
are both recognised by the host and lead to the induction of antibodies. HA functions
to allow virus attachment to sialic acid on respiratory epithelial cells followed by
membrane fusion and virus entry, while NA cleaves sialic acid, thus allowing escape
of mature virions from the infected cell [22]. Both are recognised by humoral
immunity but the induction of anti-HA antibodies in particular is well recognised to
correlate with protection and is the core mechanism by which all current influenza
vaccines function. The HA protein (which is arranged as a homotrimer) consists of a
transmembrane and two extracellular domains: a highly glycosylated and globular
head (HA1) and a stalk (HA2) that is essential for the fusogenic function of HA and
therefore relatively conserved across virus strains [23]. The HA head encompasses
the receptor binding site and a number of antigenic areas to which antibodies can
bind [24]. These antibodies may allow virus neutralisation but the sites that they
recognise are usually highly variable undergoing a process known as “antigenic drift”
[25]. Since there are relatively few functional constraints, variations in these areas
(due to the poor copy fidelity of RNA polymerase) accumulate rapidly in response to
immune pressure and gradually lead to sufficient alteration of the antigenic sites to
render antibody-mediated immunity ineffective and allow seasonal epidemics to
occur. Over time these changes have resulted in divergence of influenza A into 16
subtypes divided into two phylogenetic groups [26,27]. Thus, H1 and H5-expressing
strains, which show greater sequence relatedness, are classified together in group 1,
whilst H3 and H7 are found in group 2. NA is arranged similarly as a homotetramer
with a head, stalk and transmembrane region linked to a short cytoplasmic tail [28]. It,
too, is subject to antigenic drift and there are 9 NA subtypes in two groups of
influenza A.
Since the genome of influenza is arranged in 8 segments, HA and NA are encoded
on separate RNA strands and can re-assort independently [29]. This, along with
varied animal reservoirs and the possibility of co-infections with more than one strain
of influenza, means that new viruses can arise through genetic re-assortment in
animals with the capacity to infect humans and an array of distinct antigenic sites that
diverge from those in recently circulating seasonal strains. This phenomenon of
“antigenic shift” means that it is possible for the majority of the population to have
little pre-existing immunity against re-assorted strains, thus allowing the widespread
infection characterising pandemics. This most recently occurred in 2009 when a re-
assorted strain comprising gene segments derived from human, swine and avian
influenza viruses circulated widely and rapidly [30]. While this mostly caused a
relatively mild clinical syndrome, highly pathogenic strains including avian H5N1
continue to have the potential to cross over and cause severe disease with efficient
human-to-human transmission [31].
In addition to antibodies, there is substantial evidence that T cells play an essential
role in immunity against influenza [21]. In contrast to antibodies, T cells only
recognise viral antigens once they have been processed within the host cell. CD4+ T
cells, functioning as T follicular helper (Tfh) cells, promote the generation of optimal
antibody responses [32], while effector Th1 cells produce anti-viral cytokines and
coordinate an antiviral response. These detect foreign antigens that have been taken
up by antigen-presenting cells and digested into polypeptides for presentation in the
context of MHC class II molecules. CD8+ T cells recognise peptides derived from
viral proteins synthesised within that cell associated with MHC class I molecules and
can therefore detect and subsequently destroy virus-infected cells [33]. Thus cell-
mediated immunity is not restricted to recognition of surface glycoproteins in their
native conformation and can by induced by internal virus proteins. Since internal
proteins are often more functionally constrained, many T cell epitopes are well
conserved between strains [34]. T cells may therefore have the potential to recognise
infection by a variety of influenza viruses and escape from cell-mediated immunity by
antigenic variation occurs less frequently than from antibodies. However, the highly
diversified nature of the MHC in outbred human populations means that no single
peptide antigen can be used to induce CD4+ or CD8+ T cells in every individual and
how T cells can be effectively harnessed in vaccination is still unclear [35].
Correlates of protection against influenza
It is known that antibodies on their own can confer protection against influenza. This
has been shown in animal models in which antibodies administered parenterally or
intranasally can reduce infection rates [36,37]. Furthermore, transplacental IgG is
associated with neonatal protection [38,39] and monoclonal antibodies have
demonstrated some efficacy against experimental infection in healthy adult
volunteers [40]. Antibodies may also play a role in protection against more severe
disease [41]. These are therefore the best understood correlates of protection and a
number of assays have been developed to measure antibody titres.
The simplest assays quantify the amount of virus-binding antibodies using enzyme-
linked immunosorbent assay (ELISA) [42]. These assays benefit from being quick,
high throughput and capable of identifying the entirety of the antigen-specific
response. However, ELISA does not distinguish between antibodies of different
functionalities. The assay most commonly used as a correlate of protection is
therefore haemagglutination inhibition (HI), which quantifies antibodies that block the
agglutination of red blood cells by sialic acid on influenza virions [43,44]. HI assays
therefore act as surrogates for virus neutralisation with a standardised technique and
have been studied widely in large-scale population-based studies. Using HI,
“seroconversion” is defined as a >4-fold increase in titre whilst “seroprotection”
equates to a titre of >1:40. This definition is based on experimental infection and
epidemiological studies of mostly young adults, where a titre of 1:40 confers an
approximate 50% decrease in risk of infection. How higher HI titres relate to
protection is less clear and how they perform in prediction of infection risk in other
populations such as children and elderly adults is still debated. Nevertheless, HI titres
are considered sufficiently robust correlates of protection as to be used for licensure
of seasonal influenza vaccines via an accelerated approval process that uses
seroconversion and seroprotection as surrogate endpoints for clinical efficacy. Virus
neutralisation itself may be measured by plaque reduction neutralisation or
microneutralisation assays. These are more sensitive assays for functional
antibodies and a true measure of the ability of sera to inhibit virus attachment and
entry to cells [42]. However, these assays remain labour-intensive and reproducibility
between laboratories can be poor [45].
The role of T cells in protection against influenza in humans is less well understood.
In animal studies, models using techniques such as antibody depletion and adoptive
transfer of memory T cells have shown that both CD4+ and CD8+ T cells contribute
to optimal clearance of influenza although both may also be associated with
damaging immunopathology [46]. However, the causal relationship between
influenza-specific T cell memory and protection from severe disease in humans has
only been investigated in a limited number of studies. These have mostly consisted
of experimental human infection trials, where healthy seronegative volunteers are
infected with a known viral inoculum. These have shown negative correlations
between disease severity or viral load and pre-existing CD8+ memory T cells in one
study [47] and CD4+ memory T cells in another[48]. The single study to examine this
in a prospective cohort of naturally-infected individuals after the emergence of the
pandemic A(H1N1)2009 virus also showed a correlation between higher frequencies
of influenza-specific CD8+ memory T cells in the blood and reduced symptoms on
subsequent infection [49]. While no study in humans can ever categorically prove the
role of specific immune mechanisms in protection, these data provide compelling
evidence for the importance of T cells in influenza, with the relative contribution of
different T cell subsets probably relating to differences in study and assay design.
There remain major questions to be answered in relation to the determinants of inter-
individual variability including the influence of HLA haplotype, quality of responses
and applicability to a range of patient subgroups which limit the use of T cell assays
to predict vaccine efficacy. However, this evidence does imply that T cells can, while
not conferring sterilising immunity, provide an additional level of protection that
reduces viral load, symptoms and, therefore, risk of transmission to others.
The majority of existing data on correlates of protection against influenza rely on
measuring immune correlates in peripheral blood. This offers the advantages of ease
and reproducibility. However, with influenza being a respiratory infection primarily
involving the lung, it is increasingly clear that mucosal immunity plays an essential
role in protection both against virus entry and progression to severe disease [50–53].
While serum IgG does reach the respiratory mucosa and reduces viral load, mucosal
IgA is actively secreted in the upper and lower respiratory tract to reach substantially
higher levels than IgG especially in the nose [50]. Measuring secretory IgA remains
problematic, however, due to technical difficulties in sample collection leading to wide
variability in measurements. Recent evidence in mouse and man have also shown
the existence and importance of a recently described subset of resident memory T
(Trm) cells, which are specialised to respond early to re-encounter with pathogens at
body surfaces, detecting virus-infected cells and terminating the infection before
substantial onward spread [54]. These cells have also been implicated in innate-like
early warning functions and do not re-circulate into blood and lymphoid tissues [55]. It
remains unclear how these memory T cells relate to those found in the circulation
and whether those in blood are an independent correlate of protection or merely
reflective of those in the lung. Limitations on sampling in human studies make it
difficult to answer these questions but recent advances in the application of
experimental human influenza challenge models may enhance our knowledge of this
area.
Current vaccine strategies
Influenza vaccines became available in the 1940s, having first been made by simple
virus inactivation for intramuscular administration followed by refinement so that they
now contain well-defined quantities of HA [56]. Inactivated influenza vaccines (IIV)
are now available in high and low dose formulations with or without adjuvants and
grown in either eggs or cell culture
(http://www.cdc.gov/flu/protect/vaccine/vaccines.htm). In addition, intradermal
administration is possible, where the advantage of specialised antigen-presenting
cells in the skin may enhance immunogenicity [57]. In 2003, the live attenuated
influenza vaccine (LAIV) Flumist was licensed in the USA, comprising recombinant
viruses based on cold-adapted strains that preferentially replicate at the lower
temperatures in the nose and cannot survive at core temperature in the lung [58].
LAIV (Fluenz) was licensed in Europe in 2011 for children aged 2-18 years and it is in
this younger age group that it is most effective, with the pre-existing mucosal
immunity present in older individuals preventing virus entry and induction of an
immune response [59]. Both types of vaccine contain HA and NA from several
circulating influenza strains: two influenza A strains (H1N1 and H3N2) and one or two
B strains. Until recently, trivalent influenza vaccines were used almost exclusively but
since 2001, two lineages of influenza B (B/Victoria and the more recent B/Yamagata)
have co-circulated in many parts of the world [60]. The recognition that around half of
the trivalent vaccines in the last 10 years have been mismatched with respect to the
influenza B lineage has meant that quadrivalent vaccines are now increasingly used,
with all LAIV now quadrivalent [61]. Despite this, the highly strain-specific nature of
antibody-mediated protection against influenza means that vaccine strains must be
exactly matched to predominant circulating viruses in order to provide protection.
When vaccines strains are well matched, efficacy reaches approximately 65-85% but
vaccines in which one or more strains are mismatched have reduced efficacy of on
average 40-50%[62]. An extreme example of this was the 2014-15 season where the
bulk of clinical cases in the UK were due to influenza A/Switzerland/9715293/2013
(H3N2) while the vaccine contained A/Texas/50/2012 [63]. Early estimates
suggested vaccine effectiveness of as little as 3.4%, although these have since been
revised upwards in many settings [64].
IIV acts principally to induce systemic antibodies and has minimal capacity to
stimulate CD8+ T cells. Mucosal antibody induction is primarily limited to IgG
transudate and this is presumably sufficient to mediate the protection seen [65].
CD4+ T cells have been shown to be induced by IIV and Tfh-like cells in peripheral
blood have been found post-vaccination that preferentially help B cells to produce
anti-influenza antibodies [66]. In contrast, LAIV has the theoretical capacity to induce
both antibodies and T cells with induction of mucosal immunity (especially secretory
IgA) likely to be a major protective mechanism. Most studies of LAIV have shown
significantly less induction of systemic IgG compared with IIV, although both nasal
wash IgA and HI titres following LAIV do correlate with protection [67,68]. In addition,
LAIV-induced influenza-specific CD8+ T cells are seen albeit at low frequencies in
peripheral blood [69]. The induction of influenza-specific Trm cells by LAIV has not
yet been specifically studied. In principle, the capacity to induce local immunity in the
airway can therefore allow LAIV to reduce viral shedding by a number of
mechanisms including immune exclusion (leading to absolute prevention of
infection); early destruction of virus-infected cells to prevent progression; or reduction
in transmissible virus by antibody neutralisation, immune complex formation and
opsonisation. Whether these have a clinically significant effect on transmission is
currently unclear and there is not yet any evidence that the engagement of local
immunity confers additional advantages in this regard.
Aside from antigenic drift and shift, repeated influenza vaccination is also necessary
due to the relatively short duration of protection conferred. While influenza infection
has been shown in experimental human studies to provide protection against the
same strain for at least 7-10 years [70], the protection afforded by vaccination is
significantly shorter with an antibody half-life of less than 9 months in some studies
[71]. This is even more brief in groups such as the elderly who generally respond
more poorly to vaccination [72]. The short duration of serum antibodies can probably
be explained by limited immunogenicity of protein subunits and the attenuated nature
of LAIV, which mean that the inflammatory responses and amount of antigen are
significantly reduced compared to infection. Further understanding of how influenza
vaccines induce adaptive immune responses is therefore necessary in attempting to
improve breadth and durability of protection.
Vaccine-induced immune responses
Following intramuscular administration of IIV, antigens are taken up by antigen-
presenting cells activated by the danger signals induced by the mild tissue damage
that is caused by injection [73]. These convey the antigens to regional lymph nodes
where they present them to B cells recognising conformational epitopes and CD4+
Tfh cells. In the dark zone of lymphoid tissue, B cells undergo proliferation,
differentiation and somatic hypermutation that leads to altered B cell receptor antigen
affinity [74]. Viable daughter cells travel to the light zone where they encounter
influenza-specific Tfh cells and follicular dendritic cells, which provide the signals for
further maturation of the B cell with selection of those that express higher affinity B
cell receptors. These then travel back to the dark zone with several rounds of this
process occurring to allow affinity maturation. Most studies of the human response to
IIV have investigated B cells in the peripheral blood following vaccination. In healthy
adults, short-lived antibody-secreting cells (plasmablasts) appear after day 3 post-
vaccination and peak briefly around day 7 [75]. These can be distinguished by their
phenotypic appearance on flow cytometry and by their capacity to produce antibodies
on ex vivo re-stimulation with matched HAs in ELISpot assays. Following IIV, the
plasmablast response is dominated by HA-specific IgG-producing cells with
significantly fewer IgA-producing plasmablasts and almost none that produce IgM
[76]. This suggests that in adults the response to IIV is primarily due to recall of
memory B cells with a relatively small mucosal component. The response to IIV
contrasts with those against natural infection, where viral antigens are present for a
more protracted length of time and are associated with an inflammatory response,
which leads to a more robust and prolonged plasmablast response characterised by
a mixture of IgG- and IgA-producing cells befitting the mucosal site of infection[77].
Despite its well-documented clinical efficacy, the immune response to LAIV has been
difficult to study and no correlates of protection have been clearly characterised. In
part this is due to the relatively low magnitude of systemic responses, which are not
strongly associated with protection [67]. This implies that the protective immune
mechanisms that play the largest role are mucosal and in the nose. Following LAIV,
plasmablasts in the peripheral blood are significantly less frequent than in response
to IIV [78]. Since plasmablasts in the blood correlate with the extent of
seroconversion, this equates to lower vaccine-specific serum ELISA titres post-
vaccination. However, IgA in nasal washes negatively correlates with risk of infection
and the induction of IgA by LAIV is supported by the higher frequency of IgA-
producing plasmablasts detected. Furthermore, LAIV has been shown to induce an
early interferon-related signature that is not seen following IIV but which may
contribute to improved immunogenicity [67]. Although LAIV has been shown in some
settings to induce CD8+ T cells particularly in children, it remains unclear how much
this contributes to reduction in disease, viral shedding or heterosubtypic immunity.
Since the internal proteins of the viruses in LAIV remain identical year after year, it
might be speculated that T cells of the same specificity would be boosted by
repeated vaccinations. However, as yet, this has not been shown and there is no
evidence that repeated LAIV leads to improved breadth of protection.
Repeated annual influenza vaccination may have additional consequences, some of
which are unpredictable and possibly detrimental. For example, in a double-blind
randomised control trial of seasonal IIV, it was noted that vaccinated children were at
greater risk of virologically confirmed acute respiratory infections caused by other
pathogens [79]. This was hypothesised to be due to the absence of non-specific
protective mechanisms that an influenza infection may temporarily induce in the
respiratory tract. Certainly protection by IIV can prevent the generation of T cells by
influenza infection and this has been shown in annually vaccinated children with
cystic fibrosis who have lower frequencies of influenza-specific CD8+ T cells than
healthy unvaccinated ones while CD4+ T cell frequencies remain similar [80].
Whether this leads to clinically detectable differences in protection against newly
emergent influenza strains is again unclear. Finally, recent evidence suggests that in
individuals who are vaccinated repeatedly seasonal vaccination effectiveness is
reduced. In a longitudinal study of 7315 cases of acute respiratory illness, vaccine
efficacy against influenza A(H3N2) was significantly higher in those who had not
been vaccinated within the previous 5 years [81]. The mechanism underlying this is
uncertain but does raise the possibility of “original antigenic sin” having an impact on
the quality of vaccine-induced antibody responses. Original antigenic sin refers to the
effect of previous encounter with a similar antigen that has induced a high affinity
antibody response. B cell memory following the earlier infection or vaccination
persists and when the individual is vaccinated with a similar but different antigen, the
recall response is directed more towards the previous antigen than the current one
[82]. This might result in reduced responsiveness to the newer strain. Again, whether
these phenomena are clinically relevant in a variety of settings remains to be shown
but these studies do indicate that recurrent vaccination may have shortcomings as
well as benefits.
Improving responses to influenza vaccine
As already mentioned, the response to vaccination can be highly variable. This may
be due to immaturity of the immune system, immunosenescence or
immunosuppression but there is also wide variation even within groups with no co-
morbidities and healthy immune systems. Why these disparities occur between
individuals is only partly known in some infections [83]. Many current strategies for
improving the effectiveness of vaccines in high-risk groups have focused on
increasing the magnitude of the antibody responses to vaccination. In older adults,
administering a higher dose of HA protein has been shown to improve antibody titres
and increase protection [84]. Alternative strategies have involved the use of
adjuvants, the first of which was an oil-in-water emulsion, MF59 [85]. Including an
adjuvant in the formulation may enhance antibody induction and allow the amount of
HA protein to be reduced [86]. However, idiosyncratic severe adverse effects have
been associated with adjuvanted influenza vaccines, including recently an increased
incidence of narcolepsy in children vaccinated with Pandemrix, a monovalent
influenza A(H1N1)2009 vaccine containing ASO3 as adjuvant [87]. Further work has
also been done on improved intradermal delivery including the use of microneedles
and patches that harness the specialised antigen-presentation mechanisms in the
skin [88]. In neonates, where responses to influenza vaccination are poor, focus is
now being shifted onto maternal vaccination [39]. Recent evidence suggests that
vaccinating pregnant women in the third trimester confers approximately 50%
reduction in influenza disease to both mother and child. The relative contribution of
transplacental IgG in protecting the infant and the “cocooning” effect of reduced
transmission from a protected mother remains to be clarified [89].
Prospects for a “universal” vaccine
Although the majority of neutralising antibody responses are directed against variable
regions in the HA head, it is now recognised that antibodies are also generated in
man against the stem of HA [90]. Since the stem makes up the machinery that allows
the fusion of the viral and host cell membranes, substantial sequence divergence in
this area commonly renders the virus incompetent. The HA stem is therefore
relatively conserved. In the early 1990s, antibodies against the HA stem were
described in mice that were broadly cross-reactive and able to neutralise multiple
strains of influenza [91]. These antibodies were not detectable by HI since they did
not prevent the binding of HA to sialic acid but nevertheless neutralised infectivity by
preventing virus entry. Since then, multiple groups have shown the existence of B
cells expressing broadly cross-reactive stem-binding antibodies in humans, first in
the memory B cell compartment (by high throughput screening of immortalised
memory B cells, for example [92]) and also in acutely responding plasmablasts [90].
One monoclonal antibody has been shown to neutralise all known influenza A
subtypes and monoclonal antibodies have been developed for clinical use in passive
immunisation [93]. Broadly cross-reactive stem-binding antibodies were particularly
over-represented in the response to influenza A pandemic(H1N1)2009 infection and
monovalent vaccination [76], which led to the hypothesis that substantial alteration in
the epitopes of the HA head (such as in a re-assorted pandemic strain) allowed
memory B cell responses against the conserved HA stem to predominate. This has
been shown to also occur with vaccination against the novel HA in an H5N1 vaccine,
which supports this hypothesis [94]. Recently, several groups have constructed stem-
only vaccine candidates which have been shown to stimulate cross-reactive
antibodies and reduce the severity of disease in mice and non-human primates [95–
97].
A number of groups have focused on T cell-inducing vaccines as a method of
improving protection [98]. These are unlikely to prevent upper respiratory tract
infection itself but do have the potential to reduce the severity of disease, ideally
rendering the infection asymptomatic and minimising the release of droplets
containing large amounts of virus. T cell vaccines therefore may have a preferential
role to play in reducing transmission even when sterilising immunity is not possible.
Several vaccine candidates have been shown to induce CD8+ T cells against
influenza components. These have included recombinant adenovirus expressing NP
and M1 [99] and peptide-based vaccines based on known conserved regions of
internal influenza proteins [100]. While most have demonstrated a capacity to induce
CD8+ T cells in blood, none have clearly shown efficacy in preventing severe
disease [101]. This may be related to the compartment in which CD8+ T cells are
induced and measured. Since influenza replication occurs primarily in the respiratory
tract, local CD8+ Trm cells may be better placed and specialised to clear infection
than those with the same specificity in blood. Influenza-specific CD8+ Trm cells are
known to be more frequent in lung than spleen (in studies of donated human organ
transplant tissue) [102] and in mice, they have been shown to confer preferential
protection against influenza disease [53]. In respiratory syncytial virus (RSV) infection
of adult volunteers, these virus-specific CD8+ T cells in the airway prior to infection
correlate strongly with reduced viral load and symptoms. The extent to which
parenterally delivered T cell vaccines can induce Trm cells in the lung is unknown but
it might be hypothesised that lack of their induction could contribute to poor vaccine
efficacy.
Conclusion
Control of influenza remains a global challenge. Although mutual protection through
reducing onward transmission is a possibility, a number of hurdles exist that makes
this goal difficult to achieve. Firstly, current influenza vaccines are suboptimal with
relatively poor immunogenicity and little capacity to protect against emergent strains.
The need for annual re-vaccination, which may have immunological and clinical
consequences that are still unforeseen, is one factor in the poor vaccine uptake seen
worldwide. However, new approaches to inducing broadly cross-protective responses
are advancing and a “universal” influenza vaccine may ultimately be possible. This,
coupled with increased understanding of influenza epidemiology in different
geographical and social settings will allow better delivery of effective vaccination and
increase the likelihood of mutual protection.
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