# Neonatal and Infantile Immune Responses to Encapsulated Bacteria and Conjugate Vaccines

Authors: Peter Klein Klouwenberg; Louis Bont
Journal: Clinical and Developmental Immunology
 (2008)

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## Abstract

Encapsulated bacteria are responsible for the majority of mortality among neonates and infants. The major components on the surface of these bacteria are polysaccharides which are important virulence factors. Immunity against these components protects against disease. However, most of the polysaccharides are thymus-independent (TI)-2 antigens which induce an inadequate immune response in neonates and infants. The mechanisms that are thought to play a role in the unresponsiveness of this age group to TI-2 stimuli will be discussed. The lack of immune response may be overcome by conjugating the polysaccharides to a carrier protein. This transforms bacterial polysaccharides from a TI-2 antigen into a thymus-dependent (TD) antigen, thereby inducing an immune response and immunological memory in neonates and infants. Such conjugated vaccines have been shown to be effective against the most common causes of invasive disease caused by encapsulated bacteria in neonates and children. These and several other approaches in current vaccine development will be discussed.


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## Body

## 1. Introduction

Globally, more than 2.5 million infants die
every year from bacteremia, respiratory, and diarrhoeal diseases [1]. A limited number of
viral and bacterial pathogens are responsible for this burden of disease
among neonates and infants. A study by the WHO has identified that the most
pathogenic bacteria are encapsulated bacteria, such as Streptococcus pneumoniae, Streptococcus
pyogenes, Staphylococcus aureus, Escherichia
coli, Neisseriameningitidis, and Haemophilusinfluenza [2]. The capsule around
these pathogens is formed by a polysaccharide coating. The immunologic
advantage of this coating is evasion of phagocyte killing, as the coating
blocks complement binding and opsonization. This can be overcome by C-reactive
protein (CRP) binding [3] and the production of antibodies against
the polysaccharide [4]. This immune response confers
protection against disease. However, especially the young [5], but also the elderly
[6], have a weak immunological
response to these encapsulated bacteria due to the thymus independent (TI)
nature of these bacteria.The humoral immune response to antigens can be
divided into thymus-dependent (TD) and TI-responses [7] (see Table 1). TD
antigens consist of soluble proteins or peptides and associate with major
histocompatibility complex (MHC)
molecules at the surface of an antigen presenting cell (APC),
thus allowing the APC to interact
with CD4+ T cells. In contrast to TD antigens, TI antigens do not require T-cells
to induce an immune response [8]. Therefore, TI antigens do not or poorly induce an
immunological memory. The antibodies that are produced are primarily of the IgM
isotype and in lesser quantities IgG2 [7]. The TI antigens are
further divided into two categories based on their interaction with B cells:
type 1 (TI-1) and type 2 (TI-2) antigens [7–9]. TI-1 antigens induce proliferation and
differentiation of B lymphocytes and induce immune responses in adults, but
also in neonates [5, 10]. TI-2 antigens on the other hand induce a limited immune
response in children below two years of age, but older children and adults
react to TI-2 antigens with the formation of sufficient antibody production by
activated B-cells. TI-1 antigens include lipopolysaccharides, which is part of
the Gram-negative bacteria cell wall [6]. Therefore, immune
responses to Gram-negative bacteria are relatively sufficient in neonates and
infants compared to encapsulated bacteria, but still below levels of those of
adults [10, 11]. TI-2 antigens are bacterial
polysaccharides from encapsulated bacteria such as most S. pneumonia serotypes, N.meningitidis, and H.
influenza [12]. Infection with these
bacteria results in a reduced immunological response in neonates and therefore
they are at risk [5]. The
polysaccharides of some other bacteria such as S.pneumoniae serotype 3, however, are not classified as TI-2
antigens.Table 1
Characteristics of thymus-dependent and thymus-independent antigens
CharacteristicsThymus-dependentThymus-independent type 1Thymus-independent type 2TH-cell activation++−−IgM-IgG switch+, IgG1, IgG3+/−, IgM, IgG2 (low
quantities)−, IgMBooster response++−Immune response in neonatesHigh (but lower than
in adults)IntermediateLowDevelopment of antibody responsesAt birth3–18 m24 mExamplesProtein antigensLPSPSThe
global use of effective vaccines directed against encapsulated polysaccharide
pathogens would reduce the morbidity and mortality among newborns and infants significantly
[13]. In poor resource
countries medical care strikingly decreases after the first year of life,
necessitating the development of effective infant immunization programs [14]. The challenge for
early life immunization is to induce sustained protection circumventing
immaturity of the immune system to TI-2 antigens [15]. Neonatal and infantile antibody
responses to vaccines are of short duration and decline rapidly within a few
months [16]. This may be associated with a
resurgence of vulnerability to infection, requiring the administration of
repeated vaccinations already in the second year of life. A better
understanding of the neonatal response to polysaccharide antigens may lead to
the development of improved vaccines. This review will explore the neonatal
immune response on polysaccharide antigens and on polysaccharide protein
conjugates. Furthermore, the efficacy and drawbacks of the three main polysaccharide-conjugate
vaccines will be discussed.


## 2. Neonatal Immune Responses to Polysaccharides

There is a marked limitation in neonatal and
infantile antibody responses to most, but not all, bacterial capsular polysaccharides
[5]. The mechanisms that
are thought to account for the partial unresponsiveness of neonates to TI-2
stimuli will be reviewed in the next paragraphs.


### 2.1. B Cell Immaturity

Polysaccharide antigens localize
preferentially to the marginal-zone (MZ) B cells, found only in the spleen.
These B cells are present in low numbers at birth and the development is
deficient in neonates [17]. MZ B cells with
adult features appear after 2 years of life and coincide with the ability to
induce an immune response to
polysaccharides. Children under two years of age
have a quantitative defect in IgG2 and IgG4 isotypes [18].
Although other isotypes reach adult levels by two to three years of age, IgG2 appears
much later in development, and adult levels of this subclass are not reached
until 5–10 years of age [19]. As
the IgG2 isotype is considered as the most effective immunoglobulin against
some polysaccharides [20], the
susceptibility of neonates and infants might be due to the defect in
immunoglobulin production. Furthermore, a dysfunctional spleen or splenectomy increases the risk of
infection by encapsulated bacteria, such as S. pneumoniae and requires
antibiotic prophylaxis [21] or vaccination [22].The possibility
that B cell immaturity in the MZ B cells might cause the reduced TI-2 antigen response
was first observed in mice by Mosier et al. [23]. With the development of the murine immune system,
B cells change from IgMhiIgDlo to IgMloIgDhi and the response to TI-2 antigens coincided with the appearance of IgD, which
takes about one to two weeks in mice [24]. Furthermore, it was shown that mice with an X-linked immune
deficiency (CBA/N mice) resulting
in B cells that phenotypically resemble neonatal B-cells are unable to respond
to TI-2 antigens [23]. The murine MZ expresses the specific
intercellular adhesion molecule-grabbing nonintegrin receptor 1 (SIGNR1) which
plays a relevant role in the immune response against encapsulated bacteria [25, 26], but its human homologue has not yet been found.


### 2.2. CD21 and Complement

The immune response to polysaccharides is initiated
when polysaccharides activate complement factor C3d via the alternative pathway
[27]. The resulting
polysaccharide-complement complex subsequently localizes in MZ
B cells expressing CD21 (complement receptor 2) [27, 28]. Neonatal and infantile B cells have low
expression of CD21 which explains the inadequate response to polysaccharides [17, 28]. Interestingly, the increase of CD21
that occurs during development coincides with the response to polysaccharides.
Furthermore, neonates have relatively low levels of complement [29]. Therefore, in early
age, CD21 cannot bind the polysaccharide-C3d complex sufficiently and
antibodies are not produced. Conjugate vaccines, however, are complement
independent and induce an antibody response in neonates and young children,
which implies that MZ B cells are not needed for this response.In a murine model, Breukels et al. [30] showed that polysaccharide-conjugates
localize in the splenic MZ of neonatal mice without obvious relation to MZ B
cells. Furthermore, it was shown in adult mice that the antibody response to polysaccharides
is absent after cobra venom factor (CVF) treatment, which depletes complement.


### 2.3. T Lymphocyte and Cytokine Defects

In spite of their name, in mice TI-antigens
need the assistance of unspecific T-cells and cytokines [31]. In neonatal mice these signals are absent or
diminished and B cells activated by TI-antigens do therefore not differentiate.
Neonatal accessory cells are deficient in interleukin (IL)-1 and tumor necrosis factor (TNF)-α, which are
important for TI-2 responses [32, 33]. Supplementation of a
mouse model with these cytokines enabled neonatal B cells to clonally expand
and differentiate [6, 32]. The reduced IL-1 and -6 production in neonatal mice by macrophages
seems to be related to a decreased level of several TLR’s (TLR2, TLR4) [34, 35]. Also the increased activity of p38 MAPK, a mitogen-activated protein
kinase that is involved in the TLR intracellular signaling pathway, seems to
play a pivotal role in the unique cytokine phenotype of the neonatal
macrophages [35]. The identity of T cells involved in antibody responses to TI-2
antigens is still unknown. Kobrynski et al. [31] measured IgG antibody production after immunization
with pneumococcal polysaccharides in mice with disruptions in selected genes of
the T cell pathway. Nonclassical MHC class I-like CD1 molecules and a subset of MHC class I-dependent CD8+ cells were found to be essential for antibody responses
to TI-2 polysaccharides.


### 2.4. Other Hypotheses

Several other explanations have been proposed
and rejected. A specific B cell subset, B-1, is thought to be important for
TI-2 responses since certain TI-2 specificities are found in this subset. However,
neonates have concentrations of B1-cells comparable to adults and it seems
unlikely that this is the cause of reduced TI-2 responses [6]. Another view was
based on the increased susceptibility of immature B cells to tolerance
induction [36]. Lastly, it was
thought that TI-2 antigens cause a relative increase in suppressor T cells
compared to amplifier T cells in a murine model, which reflects an imbalance
between Th1 and Th2, in favor of Th1 cells. However, this is unsatisfactory
since the current data suggest a Th1 deficiency in neonates [6].


## 2.1. B Cell Immaturity

Polysaccharide antigens localize
preferentially to the marginal-zone (MZ) B cells, found only in the spleen.
These B cells are present in low numbers at birth and the development is
deficient in neonates [17]. MZ B cells with
adult features appear after 2 years of life and coincide with the ability to
induce an immune response to
polysaccharides. Children under two years of age
have a quantitative defect in IgG2 and IgG4 isotypes [18].
Although other isotypes reach adult levels by two to three years of age, IgG2 appears
much later in development, and adult levels of this subclass are not reached
until 5–10 years of age [19]. As
the IgG2 isotype is considered as the most effective immunoglobulin against
some polysaccharides [20], the
susceptibility of neonates and infants might be due to the defect in
immunoglobulin production. Furthermore, a dysfunctional spleen or splenectomy increases the risk of
infection by encapsulated bacteria, such as S. pneumoniae and requires
antibiotic prophylaxis [21] or vaccination [22].The possibility
that B cell immaturity in the MZ B cells might cause the reduced TI-2 antigen response
was first observed in mice by Mosier et al. [23]. With the development of the murine immune system,
B cells change from IgMhiIgDlo to IgMloIgDhi and the response to TI-2 antigens coincided with the appearance of IgD, which
takes about one to two weeks in mice [24]. Furthermore, it was shown that mice with an X-linked immune
deficiency (CBA/N mice) resulting
in B cells that phenotypically resemble neonatal B-cells are unable to respond
to TI-2 antigens [23]. The murine MZ expresses the specific
intercellular adhesion molecule-grabbing nonintegrin receptor 1 (SIGNR1) which
plays a relevant role in the immune response against encapsulated bacteria [25, 26], but its human homologue has not yet been found.


## 2.2. CD21 and Complement

The immune response to polysaccharides is initiated
when polysaccharides activate complement factor C3d via the alternative pathway
[27]. The resulting
polysaccharide-complement complex subsequently localizes in MZ
B cells expressing CD21 (complement receptor 2) [27, 28]. Neonatal and infantile B cells have low
expression of CD21 which explains the inadequate response to polysaccharides [17, 28]. Interestingly, the increase of CD21
that occurs during development coincides with the response to polysaccharides.
Furthermore, neonates have relatively low levels of complement [29]. Therefore, in early
age, CD21 cannot bind the polysaccharide-C3d complex sufficiently and
antibodies are not produced. Conjugate vaccines, however, are complement
independent and induce an antibody response in neonates and young children,
which implies that MZ B cells are not needed for this response.In a murine model, Breukels et al. [30] showed that polysaccharide-conjugates
localize in the splenic MZ of neonatal mice without obvious relation to MZ B
cells. Furthermore, it was shown in adult mice that the antibody response to polysaccharides
is absent after cobra venom factor (CVF) treatment, which depletes complement.


## 2.3. T Lymphocyte and Cytokine Defects

In spite of their name, in mice TI-antigens
need the assistance of unspecific T-cells and cytokines [31]. In neonatal mice these signals are absent or
diminished and B cells activated by TI-antigens do therefore not differentiate.
Neonatal accessory cells are deficient in interleukin (IL)-1 and tumor necrosis factor (TNF)-α, which are
important for TI-2 responses [32, 33]. Supplementation of a
mouse model with these cytokines enabled neonatal B cells to clonally expand
and differentiate [6, 32]. The reduced IL-1 and -6 production in neonatal mice by macrophages
seems to be related to a decreased level of several TLR’s (TLR2, TLR4) [34, 35]. Also the increased activity of p38 MAPK, a mitogen-activated protein
kinase that is involved in the TLR intracellular signaling pathway, seems to
play a pivotal role in the unique cytokine phenotype of the neonatal
macrophages [35]. The identity of T cells involved in antibody responses to TI-2
antigens is still unknown. Kobrynski et al. [31] measured IgG antibody production after immunization
with pneumococcal polysaccharides in mice with disruptions in selected genes of
the T cell pathway. Nonclassical MHC class I-like CD1 molecules and a subset of MHC class I-dependent CD8+ cells were found to be essential for antibody responses
to TI-2 polysaccharides.


## 2.4. Other Hypotheses

Several other explanations have been proposed
and rejected. A specific B cell subset, B-1, is thought to be important for
TI-2 responses since certain TI-2 specificities are found in this subset. However,
neonates have concentrations of B1-cells comparable to adults and it seems
unlikely that this is the cause of reduced TI-2 responses [6]. Another view was
based on the increased susceptibility of immature B cells to tolerance
induction [36]. Lastly, it was
thought that TI-2 antigens cause a relative increase in suppressor T cells
compared to amplifier T cells in a murine model, which reflects an imbalance
between Th1 and Th2, in favor of Th1 cells. However, this is unsatisfactory
since the current data suggest a Th1 deficiency in neonates [6].


## 3. Polysaccharide Conjugate Vaccines

The impaired neonatal and infantile immune
response to polysaccharide vaccines can be circumvented by conjugating the
polysaccharide to a protein carrier [37–39], based on the old dogma that haptens
when attached to a protein carrier can induce an immune response. The mechanism
by which a polysaccharide-protein conjugate vaccine acts is depicted in Figure 1. Today, all current polysaccharide vaccines registered for children below age
two are conjugated vaccines. Conjugates transform bacterial polysaccharides
from a TI-2 antigen into a TD-antigen and thereby induce an immune response and
immunological memory in neonates [40], however not to the
extend as in adults [4]. Several factors which are related
especially to immunizations might contribute to a suboptimal response to
conjugate vaccines. Firstly, the route of immunization determines the recall
response to polysaccharides in mice that have been vaccinated with a conjugate
as neonates [41]. Secondly, the choice of type of
protein conjugate is important and determines the amount of IgG antibody
response [41]. Lastly,
in neonatal mice there is a Th-2 skew which leads to a predominantly IgG1
response and impaired IgG2a antibody formation [4], the latter thought to be more
protective against encapsulated bacteria [6].Schematic diagram of mechanism of action of PS and PS-protein conjugate vaccines.
(a) Polysaccharide vaccines stimulate marginal zone B-cells which will proliferate in loco and differentiate into short-living plasma cells, which are responsible for the rapid release of low-affinity antibodies and thus first-line defence against the pathogen; (b) conjugate vaccines stimulate B-blasts that will migrate into the germinal centre, where they proliferate, undergo somatic hypermutations and isotype class switch, and differentiate into either long-living plasma cells (that produce high-affinity antibodies) and memory B cells. Abbreviations: CD40L (CD40 ligand), GC (germinal centre), MZ (marginal zone), DC (dendritic cell), MZ DC (marginal zone dendritic cell).
(a)(b)


### 3.1. Haemophilus Influenza B Conjugate Vaccine

SeveralHaemophilus
influenza B (HiB) conjugate
vaccines were introduced in the early 1990s. The reported efficacy against
meningitis and epiglottitis ranged from 94% in children of 2-3 years of age to
99% for infants below 1 year of age [42]. In Brazil, the conjugate vaccine led
to a decrease in HiB meningitis 2.39 to 0.06 cases per 100 000 population (98%)
overall, and from 60.9 to 3.1 cases per 100 000 population (95%) in children
<1 year of age, five years after the introduction of the vaccine [43]. Furthermore, the HiB conjugate vaccines
reduced carriage of HiB [44] and probably led to lower transmission
rates to children who lacked protective antibodies.


### 3.2. Neisseria Meningitidis Conjugate Vaccine

Purified polysaccharides fromN. menigitidis serogroups A, C, W135,
and Y are available vaccine products and elicit antibody responses with no
memory function, with the exception of serogroup A polysaccharide which induces
a marginal antibody response also in infants [45]. The serogroup C
polysaccharide is not immunogenic in children below 2 years of age, and
development of antibody titers is slow [46]. Conjugates have been
developed using the same principles as for HiB. The type A and C conjugate
vaccines are safe and well tolerated in infants and young children [47]. In Spain, the meningococcal
C vaccine (MCC) was effective in
98% of infants vaccinated at two, four, and six months of age and 99% in those
vaccinated after seven months of age [48]. However, the vaccine
effectiveness fell after the first year, especially in those vaccinated as
infants. In England,
the estimated effectiveness was 66% in infants vaccinated at two, three, and
four months of age and 83% in those vaccinated after seven months of age [16]. It fell to low levels after one year in
those vaccinated in the first year of life. A vaccine against type B (MenB),
which is a common cause of meningococcal disease, is presently unavailable [49, 50]. However, unpublished data
provide a source of optimism to develop a safe and effective containing
recombinant outer membrane surface proteins of MenB vaccine from the N.
meningitidis strain NZ98/254. This investigational vaccine against MenB
induced protective immune responses against 3 strains after the third dose
(Miller et al. abstract 133, Annual Meeting of the European Society of
Paediatric Infectious Diseases (ESPID, 2008)). Results from Phase 3 RCTs are
expected during the coming years.


### 3.3. Streptococcus Pneumoniae Conjugate Vaccine

Diseases caused by pneumococci include
pneumonia, meningitis, otitis media, sinusitis, and bronchitis. Pneumococcal vaccines
that are effective in infants have been welcomed as resistance against growing commonly
used antibiotics [51]. An unconjugated 23-valent vaccine is registered
for children over two years of age but is ineffective in younger children [52]. A 7-valent
polysaccharide-protein conjugate vaccine (PCV-7) has been introduced for use in
children below the age of 2 years. Serotypes included in PCV-7 cover 65–80% of serotypes
that cause invasive pneumococcal infections [53]. Other conjugate vaccines with wider
serotype coverage, including a 10-valent vaccine and a 13-valent vaccine, are
in the late stage of development [54, 55]. A
94% decrease in vaccine type invasive pneumococcal disease in children with a
coverage rate of just 68% was reported [56]. The number of
all-cause pneumonia admission rates has declined by 39% for children younger
than two years in the USA from 2000–2004 [57] and admissions due to pneumococcal
meningitis were reduced by 66% in the same period [58]. A significant
decrease was also seen in the unvaccinated groups as a result of herd immunity
due to decreased transmission from vaccinated children to unvaccinated contacts
[56]. The protective efficacy against acute
otitis media however has been relatively modest. In a Finnish study, the
efficacy against confirmed otitis media was 34% and the overall efficacy
against otitis media regardless of cause was only 6% [59].


### 3.4. Other Conjugate Vaccines

Several other polysaccharide pathogens are
under investigation. Group B streptococci are the major cause of meningitis and
sepsis in neonates. In animal studies, group B streptococcal conjugate vaccines
have shown to be able to induce protective antibodies [60]. Similar attempts have been made to
develop immunogenic and safe vaccines against the Vi polysaccharide of S. typhi [61] and polysaccharides and LPS of E. coli [62] as well as S. aureus [63].


## 3.1. Haemophilus Influenza B Conjugate Vaccine

SeveralHaemophilus
influenza B (HiB) conjugate
vaccines were introduced in the early 1990s. The reported efficacy against
meningitis and epiglottitis ranged from 94% in children of 2-3 years of age to
99% for infants below 1 year of age [42]. In Brazil, the conjugate vaccine led
to a decrease in HiB meningitis 2.39 to 0.06 cases per 100 000 population (98%)
overall, and from 60.9 to 3.1 cases per 100 000 population (95%) in children
<1 year of age, five years after the introduction of the vaccine [43]. Furthermore, the HiB conjugate vaccines
reduced carriage of HiB [44] and probably led to lower transmission
rates to children who lacked protective antibodies.


## 3.2. Neisseria Meningitidis Conjugate Vaccine

Purified polysaccharides fromN. menigitidis serogroups A, C, W135,
and Y are available vaccine products and elicit antibody responses with no
memory function, with the exception of serogroup A polysaccharide which induces
a marginal antibody response also in infants [45]. The serogroup C
polysaccharide is not immunogenic in children below 2 years of age, and
development of antibody titers is slow [46]. Conjugates have been
developed using the same principles as for HiB. The type A and C conjugate
vaccines are safe and well tolerated in infants and young children [47]. In Spain, the meningococcal
C vaccine (MCC) was effective in
98% of infants vaccinated at two, four, and six months of age and 99% in those
vaccinated after seven months of age [48]. However, the vaccine
effectiveness fell after the first year, especially in those vaccinated as
infants. In England,
the estimated effectiveness was 66% in infants vaccinated at two, three, and
four months of age and 83% in those vaccinated after seven months of age [16]. It fell to low levels after one year in
those vaccinated in the first year of life. A vaccine against type B (MenB),
which is a common cause of meningococcal disease, is presently unavailable [49, 50]. However, unpublished data
provide a source of optimism to develop a safe and effective containing
recombinant outer membrane surface proteins of MenB vaccine from the N.
meningitidis strain NZ98/254. This investigational vaccine against MenB
induced protective immune responses against 3 strains after the third dose
(Miller et al. abstract 133, Annual Meeting of the European Society of
Paediatric Infectious Diseases (ESPID, 2008)). Results from Phase 3 RCTs are
expected during the coming years.


## 3.3. Streptococcus Pneumoniae Conjugate Vaccine

Diseases caused by pneumococci include
pneumonia, meningitis, otitis media, sinusitis, and bronchitis. Pneumococcal vaccines
that are effective in infants have been welcomed as resistance against growing commonly
used antibiotics [51]. An unconjugated 23-valent vaccine is registered
for children over two years of age but is ineffective in younger children [52]. A 7-valent
polysaccharide-protein conjugate vaccine (PCV-7) has been introduced for use in
children below the age of 2 years. Serotypes included in PCV-7 cover 65–80% of serotypes
that cause invasive pneumococcal infections [53]. Other conjugate vaccines with wider
serotype coverage, including a 10-valent vaccine and a 13-valent vaccine, are
in the late stage of development [54, 55]. A
94% decrease in vaccine type invasive pneumococcal disease in children with a
coverage rate of just 68% was reported [56]. The number of
all-cause pneumonia admission rates has declined by 39% for children younger
than two years in the USA from 2000–2004 [57] and admissions due to pneumococcal
meningitis were reduced by 66% in the same period [58]. A significant
decrease was also seen in the unvaccinated groups as a result of herd immunity
due to decreased transmission from vaccinated children to unvaccinated contacts
[56]. The protective efficacy against acute
otitis media however has been relatively modest. In a Finnish study, the
efficacy against confirmed otitis media was 34% and the overall efficacy
against otitis media regardless of cause was only 6% [59].


## 3.4. Other Conjugate Vaccines

Several other polysaccharide pathogens are
under investigation. Group B streptococci are the major cause of meningitis and
sepsis in neonates. In animal studies, group B streptococcal conjugate vaccines
have shown to be able to induce protective antibodies [60]. Similar attempts have been made to
develop immunogenic and safe vaccines against the Vi polysaccharide of S. typhi [61] and polysaccharides and LPS of E. coli [62] as well as S. aureus [63].


## 4. Drawbacks of Conjugate Vaccines

As described above, polysaccharide-conjugate
vaccines are effective in children under the age of two years. However,
conjugate vaccines are only available forH.
influenza type B, some meningococcal subtypes, and recently for a limited
number of subtypes of S. pneumoniae. Vaccine
failure and rise of nonserotype bacteria have been noticed and require
continuous attention [64]. There is evidence that coadministration
with other vaccines may impair effectiveness of the vaccinations [65]. Finally, conjugate vaccines are
expensive, which does not allow for global use of these vaccines [53].


### 4.1. Haemophilus Influenza B Conjugate Vaccine

A decade after its introduction in the UK, the
first conjugate vaccine, HiB, became less effective and vaccine failures were
seen [66]. Two explanations
have been given for this effect. First, after early age immunization antibody
levels are sufficient for protection but drop over the following years,
sometimes to levels that are not considered protective. This was explained by a
reduction
in either the number or the quality of memory B cells induced by immunization
or a loss of avidity in matured B cells prior to disease onset following
defective priming [67]. The low titers
observed in the UK
may have been exacerbated by the loss of “natural
boosting” associated with a reduction in carriage. Reboosting might be the
solution [67]. The other important factor contributing
to the reduced immune response to HiB-conjugate-vaccine is the current
coadministration with other vaccines. Reduced antibody responses to HiB
conjugates have been documented using acellular pertussis/HiB [65], DTaP-HiB [68], and MCC/HiB combinations [69], but not in other studies [70, 71]. The impaired immune responses were
increased by accelerated immunization schedules such as that in the UK
[72]. The precise immunological mechanism
responsible for the excess of vaccine failures following the combination
vaccine is not known.


### 4.2. Neisseria Meningitidis Conjugate Vaccine

The meningococcal serogroup B polysaccharide is
poorly immunogenic in man [73]. The development of an effective vaccine
against N. meningitidis serogroup B
is complicated by the inability of this polysaccharide to induce a significant antibody
response [73], even when conjugated to a carrier
protein [74]. However, unpublished data show
promising results (Miller et al. abstract 133, Annual Meeting of the
European Society of Paediatric Infectious Diseases (ESPID, 2008)). In Spain and the UK,
the MCC vaccine effectiveness fell
after the first year, especially in those vaccinated as infants [16, 48]. These data suggest that the protection
given by MCC vaccine may be age dependent
and that children vaccinated at an older age may have greater and
longer-lasting protection than those vaccinated as infants. This suggests that
protection may be more reliant on circulating antibodies at the time of
exposure than on the ability to mount a booster response [75]. A recent study found that the immune response and length of protection were dependent on the formation of a large germinal center one month after
primary immunization with the MCC vaccine [76]. One third of infants in
this study produced a very low number of memory B cells after the initial
immunization and did not maintain protective antibody levels by one year of
age. In these children, the germinal center was underdeveloped. Understanding
of the factors that determine the production of these germinal centers could
lead to improved conjugate vaccines. Until then, booster doses of MCC may be required in order to extend the duration
of protection offered by the vaccine.


### 4.3. Streptococcus Pneumoniae Conjugate Vaccine

A major drawback of pneumococcal conjugate
vaccines is that the serotypes included in PCV-7 cover only 65–80% of serotypes
that cause invasive pneumococcal infections [53]. Efforts to include more subtypes in a
conjugate vaccine prove to be very complicated and costly. Furthermore, it has
been shown that combination of PCV-7 with other vaccines can lead to reduced immune
responses. The response to hepatitis B vaccine was nonsignificantly reduced
with concomitant administration with PCV-7 [77]. Another problem is that the PCV-7
vaccine replaces disease by nonvaccine serotypes especially 19A [78] and 16F [59]. A recent study in Alaska, where routine vaccination in
children has started in 1999–2000, showed an
increase in invasive pneumococcal disease rate caused by nonvaccine serotypes
of 140% compared with the prevaccine period [64]. In the first three years after
introduction of the PCV-7 vaccine, there was a 96% decrease in heptavalent
vaccine serotype disease. This led to a decrease in overall invasive
pneumococcal disease of 67% in Alaskan children younger than 2 years (from 403.2
per 100 000 in 1995–2000 to 134.3 per
100 000 per year in 2001–2003) but to an
82% increase in invasive disease in the following years to 244.6/100 000.
Serotype 19A accounted for 28% of invasive pneumococcal disease among Alaska
children younger
than 2 years during 2004–2006. There was
no significant increase in disease due to nonvaccine serotypes in nonnative
Alaskan children younger than 2 years [64]. This emphasizes the importance of
continuing surveillance and development of expanded valency vaccines. The
question remains whether this serotype shift leads to increased morbidity and
mortality rates as these nonvaccine types are typically less pathogenic. Other
limitations are a modest effect on nasopharyngeal colonization [79], cost (US$ 32 000–166 000 per life-year
saved) [53], and difficulties in production that
have led to shortages.


## 4.1. Haemophilus Influenza B Conjugate Vaccine

A decade after its introduction in the UK, the
first conjugate vaccine, HiB, became less effective and vaccine failures were
seen [66]. Two explanations
have been given for this effect. First, after early age immunization antibody
levels are sufficient for protection but drop over the following years,
sometimes to levels that are not considered protective. This was explained by a
reduction
in either the number or the quality of memory B cells induced by immunization
or a loss of avidity in matured B cells prior to disease onset following
defective priming [67]. The low titers
observed in the UK
may have been exacerbated by the loss of “natural
boosting” associated with a reduction in carriage. Reboosting might be the
solution [67]. The other important factor contributing
to the reduced immune response to HiB-conjugate-vaccine is the current
coadministration with other vaccines. Reduced antibody responses to HiB
conjugates have been documented using acellular pertussis/HiB [65], DTaP-HiB [68], and MCC/HiB combinations [69], but not in other studies [70, 71]. The impaired immune responses were
increased by accelerated immunization schedules such as that in the UK
[72]. The precise immunological mechanism
responsible for the excess of vaccine failures following the combination
vaccine is not known.


## 4.2. Neisseria Meningitidis Conjugate Vaccine

The meningococcal serogroup B polysaccharide is
poorly immunogenic in man [73]. The development of an effective vaccine
against N. meningitidis serogroup B
is complicated by the inability of this polysaccharide to induce a significant antibody
response [73], even when conjugated to a carrier
protein [74]. However, unpublished data show
promising results (Miller et al. abstract 133, Annual Meeting of the
European Society of Paediatric Infectious Diseases (ESPID, 2008)). In Spain and the UK,
the MCC vaccine effectiveness fell
after the first year, especially in those vaccinated as infants [16, 48]. These data suggest that the protection
given by MCC vaccine may be age dependent
and that children vaccinated at an older age may have greater and
longer-lasting protection than those vaccinated as infants. This suggests that
protection may be more reliant on circulating antibodies at the time of
exposure than on the ability to mount a booster response [75]. A recent study found that the immune response and length of protection were dependent on the formation of a large germinal center one month after
primary immunization with the MCC vaccine [76]. One third of infants in
this study produced a very low number of memory B cells after the initial
immunization and did not maintain protective antibody levels by one year of
age. In these children, the germinal center was underdeveloped. Understanding
of the factors that determine the production of these germinal centers could
lead to improved conjugate vaccines. Until then, booster doses of MCC may be required in order to extend the duration
of protection offered by the vaccine.


## 4.3. Streptococcus Pneumoniae Conjugate Vaccine

A major drawback of pneumococcal conjugate
vaccines is that the serotypes included in PCV-7 cover only 65–80% of serotypes
that cause invasive pneumococcal infections [53]. Efforts to include more subtypes in a
conjugate vaccine prove to be very complicated and costly. Furthermore, it has
been shown that combination of PCV-7 with other vaccines can lead to reduced immune
responses. The response to hepatitis B vaccine was nonsignificantly reduced
with concomitant administration with PCV-7 [77]. Another problem is that the PCV-7
vaccine replaces disease by nonvaccine serotypes especially 19A [78] and 16F [59]. A recent study in Alaska, where routine vaccination in
children has started in 1999–2000, showed an
increase in invasive pneumococcal disease rate caused by nonvaccine serotypes
of 140% compared with the prevaccine period [64]. In the first three years after
introduction of the PCV-7 vaccine, there was a 96% decrease in heptavalent
vaccine serotype disease. This led to a decrease in overall invasive
pneumococcal disease of 67% in Alaskan children younger than 2 years (from 403.2
per 100 000 in 1995–2000 to 134.3 per
100 000 per year in 2001–2003) but to an
82% increase in invasive disease in the following years to 244.6/100 000.
Serotype 19A accounted for 28% of invasive pneumococcal disease among Alaska
children younger
than 2 years during 2004–2006. There was
no significant increase in disease due to nonvaccine serotypes in nonnative
Alaskan children younger than 2 years [64]. This emphasizes the importance of
continuing surveillance and development of expanded valency vaccines. The
question remains whether this serotype shift leads to increased morbidity and
mortality rates as these nonvaccine types are typically less pathogenic. Other
limitations are a modest effect on nasopharyngeal colonization [79], cost (US$ 32 000–166 000 per life-year
saved) [53], and difficulties in production that
have led to shortages.


## 5. New Developments

The currently registered conjugated polysaccharide
vaccines have been developed based upon the principle that CD4 T cell
recruitment is necessary for the activation of the infant B cell immune response
[8]. The underlying
thought was to promote a transformation of the neonatal immune response from a TI
one to a TD one by conjugating the polysaccharides to immunogenic carrier
proteins. The neonatal immune response to TD-antigens has been shown to be
better than the response to TI-2 antigens but did not reach levels seen in
healthy adults [4]. One of the most important factors determining
the neonatal immune response to conjugate vaccines seems to the type of carrier
protein used in the conjugate [4]. The carrier determines the level of induction
of specific T cells and therefore the levels of polysaccharide antibodies and
hence the protective effect gained by administration of the conjugate vaccine. The
optimal carrier might be different for different pneumococcal serotypes [80]. In one study pneumococcal
polysaccharide conjugated to a diphtheria carrier was more efficient in
inducing a mucosal response, while tetanus conjugate resulted in improved
systemic responses [81]. Another study showed that a tetanus
conjugate resulted in a better serotype 4 response, while a diphtheria
conjugate evoked a better response to types 3, 9 V, and 14 [82].The addition of adjuvants to conjugate
vaccines could potentially reduce the number of doses needed to establish
protective immunity and thereby provides protective immunity within a shorter time period and at
a reduced cost. Adjuvants also lead to more consistent induction of responses
to various polysaccharide serotypes [83]. Toll-like receptor (TLR) ligands have been considered as
vaccine adjuvants [84, 85], such as CpG containing oligodeoxynucleotides [86]. Peptide p458 is a peptide derived from the human or mouse 60-kDa heat shock protein
(hsp60) and stimulates TLR4 [84]. Conjugated to
pneumococcal polysaccharide type 4 it could induce protection in mice against a
supralethal S. pneumoniae challenge.
Protection was associated with polysaccharide type 4-specific IgG antibodies in
most but not in all the mice, a T cell response to the p458 carrier and
long-term memory. Vaccines composed of p458 conjugated to the polysaccharides
of Salmonella [87], or
meningococcus B and C [88] were also
immunogenic in mice, even when injected without an added adjuvant. Other TLR
agonists that stimulate TLR8, such as R-848 (TLR7/8), the imidazoquinoline
congeners 3M-003 (TLR7/8) and 3M-002 (TLR8), as well as single-stranded viral
RNAs (TLR8), also induce a strong immune response in neonates and infants by
stimulating p38 MAPK phosphorylation [85]. Furthermore, LT-K63, a nontoxic mutant of E. coli heat-labile enterotoxin [89], when administered concomitantly with a
conjugated pneumococcal polysaccharide serotype 1, enhanced IgG responses in infant
mice compared to conjugated polysaccharide alone [89]. A second dose of conjugated
pneumococcal polysaccharide resulted in very high IgG responses and
significantly improved protection against lethal pneumococcal infections in
this animal model. Similar results were obtained with an MCC vaccine [90].Furthermore, the route of administration is
also shown to be an important determinant in eliciting protective immunity in
neonates [4, 89]. Intranasal immunization with conjugated
polysaccharides [89, 91] seemed to be effective both in infantile
and neonatal mice. A single intranasal dose of conjugate vaccine elicited a
sufficient high IgG response to protect neonatal mice against pneumococcal
infections, whereas subcutaneous administration required two doses to induce
complete protection. The increased efficiency could be explained by the additional
induction of a salivary IgA response after intranasal administration. However,
antibody responses and protective efficacy remained significantly lower than in
adult mice [89]. One of the main reasons for this seems
to be the lack of effective adjuvants [92].Another strategy in early stage of development
is the use of surface proteins. An example is the Pneumococcal surface protein
A (PspA) that is a cell-wall-associated surface protein [93]. It is known to play a major role in the
pneumococcal virulence; it binds human lactoferrin and interferes with
complement deposition on the bacterial surface. It is thought that it might
result in better immune responses in infants and neonates. The antibody
response to PspA has been studied in children [93, 94]. The pneumococcal surface antigen
protein A (PsaA) is currently explored as a vaccine candidate. It is
structurally conserved [95] and plays a role in adherence to host
mucosae [96]. Until now, however,
it has not been used as vaccine antigen in humans.Another potential vaccine strategy is the
development of peptides that mimic polysaccharide antigens [97]. The main advantage
of using peptides over polysaccharides is that peptides induce a TD antigen
response as they are processed by APC’s
and presented to T cells. A drawback of the use of peptides in vaccines is
their poor chemical stability and subsequently lower immunogenicity in vivo. DNA-based vaccines are another potential approach
as they are more stable but were initially not considered a viable option for
pathogens coated with polysaccharides since carbohydrate antigens are secondary
gene products [97]. However, it was
recently shown that a DNAvaccine
could induce an IgG2a isotype response against a polysaccharide antigen [98]. Other possible advantages of DNA-vaccines are the relatively straightforward and
cheaper production techniques compared to conjugate vaccines.


## 6. Conclusion

Neonatal immune responses to polysaccharide
pathogens are very weak. Therefore, neonates and young children are at risk for
invasive infections withS. pneumococcus, N. meningitidis, and H. influenza. An important percentage of
deaths among neonates are caused by these bacteria. The efficacy of the currently
used conjugate vaccines is already very high in the population most at risk,
but worldwide utilization of these vaccines is hampered by high production
costs. Knowledge about neonatal immunological responses to polysaccharide
antigens may open the way for the application of newly designed conjugated
vaccines or vaccines based on other principles in this patient group.
Currently, several strategies are being explored to get insight into the
mechanisms underlying the limitations of infant responses and to thereby improve
neonatal vaccination efficiency.



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*Source: 628963-2008-09-23.xml*