Since January 2020 Elsevier has created a COVID-19 resource centre with

free information in English and Mandarin on the novel coronavirus COVID-

19. The COVID-19 resource centre is hosted on Elsevier Connect, the
company's public news and information website.

Elsevier hereby grants permission to make all its COVID-19-related

research that is available on the COVID-19 resource centre - including this
research content - immediately available in PubMed Central and other
publicly funded repositories, such as the WHO COVID database with rights
for unrestricted research re-use and analyses in any form or by any means
with acknowledgement of the original source. These permissions are
granted for free by Elsevier for as long as the COVID-19 resource centre
remains active.
 Phytomedicine 60 (2019) 152905

Contents lists available at ScienceDirect

Phytomedicine
journal homepage: www.elsevier.com/locate/phymed

Original Article

Updated insights into the mechanism of action and clinical profile of the T
immunoadjuvant QS-21: A review☆
Marie-Aleth Lacaille-Dubois
PEPITE EA 4267, Université de Bourgogne Franche-Comté, Laboratoire de Pharmacognosie, UFR des Sciences de Santé, 7, Bd Jeanne d'Arc, 21079 Dijon Cedex, France

ARTICLE INFO ABSTRACT

Keywords: Background: Vaccine adjuvants are compounds that significantly enhance/prolong the immune response to a co-
Quillaja saponaria administered antigen. The limitations of the use of aluminium salts that are unable to elicite cell responses
QS-21 against intracellular pathogens such as those causing malaria, tuberculosis, or AIDS, have driven the develop-
Vaccine adjuvant ment of new alternative adjuvants such as QS-21, a triterpene saponin purified from Quillaja saponaria.
Herpes zoster
Purpose: The aim of this review is to attempt to clarify the mechanism of action of QS-21 through either re-
Malaria
ceptors or signaling pathways in vitro and in vivo with special emphasis on the co-administration with other
Cancer
immunostimulants in new adjuvant formulations, called adjuvant systems (AS). Furthermore, the most relevant
clinical applications will be presented.
Methods: A literature search covering the period 2014–2018 was performed using electronic databases from Sci
finder, Science direct, Medline/Pubmed, Scopus, Google scholar.
Results: Insights into the mechanism of action of QS-21 can be summarized as follows: 1) in vivo stimulation of
Th2 humoral and Th1 cell-mediated immune responses through action on antigen presenting cells (APCs) and T
cells, leading to release of Th1 cytokines participating in the elimination of intracellular pathogens. 2) activation
of the NLRP3 inflammasome in mouse APCs with subsequent release of caspase-1 dependent cytokines, Il-1β and
Il-18, important for Th1 responses. 3) synthesis of nearly 50 QS-21 analogs, allowing structure/activity re-
lationships and mechanistic studies. 4) unique synergy mechanism between monophosphoryl lipid A (MPL A)
and QS-21, formulated in a liposome (AS01) in the early IFN-γ response, promoting vaccine immunogenicity.
The second part of the review is related to phase I-III clinical trials of QS-21, mostly formulated in ASs, to
evaluate efficacy, immunogenicity and safety of adjuvanted prophylactic vaccines against infectious diseases,
e.g. malaria, herpes zoster, tuberculosis, AIDS and therapeutic vaccines against cancer and Alzheimer's disease.
Conclusion: The most advanced phase III clinical applications led to the development of two vaccines containing
QS-21 as part of the AS, the Herpes Zoster vaccine (HZ/su) (Shingrix™) which received a license in 2017 from the
FDA and a marketing authorization in the EU in 2018 and the RTS,S/AS01 vaccine (Mosquirix™) against malaria,
which was approved by the EMA in 2015 for further implementation in Sub-Saharan countries for routine use.

Introduction
Vaccines are the most effective and least expensive methodology for
preventing diseases caused by infectious pathogens. Recently emerging
or re-emerging diseases such as the severe acute respiratory syndrome
(SARS) in 2003, H1N1 influenza pandemic in 2009 or Ebola virus in
2014 have made the research on vaccines very attractive in the last
decades (Lee and Nguyen, 2015). Modern subunit vaccines comprising
homogeneous molecular antigens have been developed to prevent and
treat many human diseases, but they are weakly immunogenic and
must be administered with an adjuvant to elicit a potent immune re-
sponse. Adjuvants whose name originates from the Latin word adjuvare

Abbreviations: A-β, amyloid-beta; APCs, antigen presenting cells; CD, cluster of differentiation; CMI, cell mediated immunity; CTL, cytotoxic T lymphocytes; DCs,
dendritic cells; dLN, draining lymph nodes; HIV, human immunodeficiency virus; HZ, herpes zoster; IFN-γ, interferon-gamma; Il-2, interleukine 2; KLH, keyhole
limpet hemocyanin; MAPK, mitogen activated protein kinase; MHC, major histocompatibility complex; MPL, 3-deacylated monophosphoryl lipid; Mtb,
Mycobacterium tuberculosis bacteria; NK, natural killer; NSCLC, non small cell lung carcinoma; PAMPs, pathogen-associated molecular patterns; PRRs, pathogen
recognition receptors; QS-21, Quillaja saponaria Molina-fraction 21; SARS, severe acute respiratory syndrome; TCR, T-cell receptor; TLR, Toll-like receptor; TNF-α,
tumor necrosis factor-alpha; VZV, varicella zoster virus
☆
Dedicated to Pr. Dr. Dr. h.c. mult. em. Hildebert Wagner in the occasion of his 90th birthday in 2019.
E-mail address: m-a.lacaille-dubois@u-bourgogne.fr.

https://doi.org/10.1016/j.phymed.2019.152905
Received 28 January 2019; Received in revised form 26 March 2019; Accepted 30 March 2019
0944-7113/ © 2019 Elsevier GmbH. All rights reserved.
M.-A. Lacaille-Dubois
Fig. 1. Schematic representation of the mechanism of action of an adjuvanted vaccine (adapted from Awate et al., 2013; Reed et al., 2013; Lacaille-Dubois and
Wagner, 2017).
(meaning to help) are substances used in combination with a specific
antigen (Ag) to produce a more potent and persistent immune response
against a specific disease than the Ag alone. The aim is to confer long-
term protection, and with the additional benefit that less antigen and
fewer injections are needed (Lee and Nguyen, 2015). Adjuvants are
used in many vaccines, but their mechanisms of action have not been
fully elucidated. They include diverse classes of compounds such as
mineral salts, microbial products, emulsions, saponins, cytokines,
polymers, microparticules and liposomes (Awate et al., 2013).
Up to now, few adjuvants have been approved for use in humans
(Del Giudice et al., 2018). Aluminium salts, which have been ex-
tensively used as adjuvants in vaccines for more than 80 years, induced
robust antibody responses but weak Th1 cell type responses, which are
instrumental for protection against many pathogens. The new genera-
tion of vaccines using purer components for safer vaccines (e.g.,subunits
vaccines with highly purified recombinant antigens) are less im-
munogenic in contrast to live attenuated or inactivated whole-cell
vaccines. Therefore, they required the induction of strong cellular re-
sponses including CD4+ T-helper cells (Th) and CD8+ cytotoxic T
lymphocytes (CTL) in addition to antibody (Ab) responses. Thus, the
development of new adjuvants is necessary for vaccines that require a
cell-mediated immune response (CMI). Among them, the triterpene
glycosides represent an interesting class of clinically relevant adjuvants,
particularly those of Quillaja saponaria (QS). These amphiphatic plant
glycosides possess a variety of pharmacological activities including
immunoadjuvant, antitumor, anti-inflammatory, and antimicrobial
properties which have been extensively studied in many in vitro and in
vivo bioassays (Lacaille-Dubois and Wagner, 1996, 2000, 2017;
Lacaille-Dubois, 1999, 2005). Despite the apparent success of QS-21 as
vaccine-adjuvant in the last years, there is an urgent need for a deeper
understanding of its mode of action which could accelerate the devel-
opment of new vaccine strategies. This requires an interdisciplinary
approach between chemists, biochemists, molecular biologists and im-
munologists. The following review will summarize updated insights
into the modes of action, the structure/activity relationships allowing
detailed mechanistic studies, and the clinical status of the vaccine ad-
juvant QS-21.
2
Phytomedicine 60 (2019) 152905
General mechanism of action of adjuvants
With a growing understanding of the innate immune system (re-
sponse first mediated by antigen presenting cells (APCs)) and its role in
activation and modulation of adaptive immune responses (Ag- specific
B and T cell responses) (Didierlaurent et al., 2017; Di Pasquale et al.,
2015), the mechanisms of action of adjuvants are being elucidated.
They can act on one or more of the following targets to increase re-
sponse to Ags: (1) sustaining release at the injection site (depot effect),
(2) transient secretion of cytokines and chemokines, (3) recruitement of
various immune cells (neutrophils, monocytes, eosinophils, macro-
phages and Dendritic Cells (DCs) at the injection site leading to a local
immune-competent environment, (4) expression by the recruited APCs
of various Pathogen Recognition Receptors (PRRs) both on their surface
(Toll-like receptors, TLRs, C-type lectin receptors, CLRs), and in-
tracellularly (Nucleotide Oligomerization Domain (NOD)-like receptors
(NLRs) and Retinoic Inducible Gene-1 (RIG)-like receptors (RLRs)),
which are recognized and/or activated by adjuvants, (5) maturation
and activation of recruited APCs which up-regulate the expression of
Major Histocompatibility Complex (MHC)-I and/or MHC-II and acti-
vation of co-stimulatory signals CD40, CD80/86, (6) increased capacity
of APCs for Ag processing and presentation by MHC, (7) migration of
the mature APCs to the draining lymph nodes (dLNs) to interact with
Ag-specific B or T lymphocytes (through receptor-ligand interactions,
MHC-T cell receptor (MHC-TCR), CD40-CD40L, CD80/86-CD28) which
are activated to produce potent Ab-secreting B cells and/or effector
CD8+ T cell responses (Awate et al., 2013).
Researchers continue to characterize the adaptive immune response
and to clarify the respective roles of vaccine-induced immune effectors
(Fig. 1). They include Abs produced by B lymphocytes that bind spe-
cifically to a toxin or a pathogen, memory B cells produced only when B
cells received T cell help having the ability to respond rapidely on re-
exposure of the same antigen, cytotoxic CD8+ T lymphocytes that limit
the spread of infectious agents by killing infected cells or secreting
specific cytokines, and CD4+ T helper (Th) lymphocytes. Various Th
subsets have been defined through their cytokine profiles and ability to
induce B cell and CD8+ T cell responses (Reed et al., 2013;
Siegrist 2013). The Th1-type CD4+T cells essentially release the
M.-A. Lacaille-Dubois
proinflammatory cytokines interleukin-2 (Il-2), interferon-γ (IFN-γ),
tumor necrosis factor-α (TNF-α), which participate in the elimination of
intracellular pathogens both directly (cytokine response) or indirectly
via CD8+ T cell activation and differentiation in CTLs (cellular immune
response) and the generation in mice of the complement-fixing Abs
IgG2a and IgG3. The Th2-type CD4+ T cell response is characterized by
secretion of Il-4, Il-5 and Il-6 providing B cell help and the preferential
induction of IgG1, IgE, and IgA in mice, and IgE and IgG4 in humans
(humoral response) (Guy, 2007).
Structure of QS-21 and proposed immunoadjuvant mechanism of
action
The saponin QS-21 has been isolated from Quillaja saponaria tree
bark (Kensil et al., 1991) and is one of the most potent adjuvants known
which is currently used in exploratory, and a few licensed, vaccines. QS-
21 is a mixture of two isomeric molecules, QS-21 Apiose (QS-21 Api),
and QS-21 Xylose (QS-21-Xyl) (2:1), each having four domains: the
triterpene quillaic acid, a branched trisaccharide linked at C-3 through
an O-heterosidic bound, a linear tetrasaccharide linked at C-28 through
an ester bound, and a glycosylated pseudodimeric acyl chain attached
through a labile ester linkage at C-4 of the fucose unit of the tetra-
saccharide (Fig. 2). This compound showed a potent adjuvant activity
against a wide variety of Ags with low toxicity in preclinical studies in
mice, guinea pigs, monkeys and baboons. It was shown to stimulate Th1
and Th2 immune responses, when added to vaccine (Kensil, 1996). This
adjuvant is a candidate for many clinical trials of vaccines directed
against infectious diseases, cancer, and others. However, the exact role
of QS-21, either through activation of receptors or signaling pathways,
is poorly characterized. Some hypotheses advanced that QS-21 may
facilitate Ag uptake into APCs by binding to cell surface lectins through
its carbohydrate domains, leading to specific cytokine profiles that
enhance the T and/or B cells’ response (Marciani, 2003). An effective
mechanism of action in which QS-21 apparently acts on both DCs and T
cells was proposed (Marciani, 2018) (Fig. 3). Exogenous Ags and QS-21
enter DCs by cholesterol-dependent endocytosis. The high affinity of
QS-21 for endosomal membrane cholesterol resulted in the destabili-
zation of the membrane (Lorent et al., 2014) leading to pore formation,
thus facilitating the escape of the antigen to the cytosol for further
processing inside the cell into peptides. They are then loaded to MHC-I
and presented on the DC surface to naïve CD8+ T cells to yield CTLs.
The role of aldehyde in QS-21 was highlighted in the T cell by forming
an imine with the amino group of the T cell surface receptor such as
CD2, providing a costimulatory signal to the T cell. This signal together
with the mitogen-activated protein kinase (MAPK) ERK2 resulting from
Fig. 2. Structure of QS-21. Glc A: β-D-glucuronopyranosyl; Glc : β-D-glucopyranosyl; Gal : β-D-galactopyranosyl; Xyl : β-D-xylopyranosyl; Fuc: β-D-fucopyranosyl;
Api: β-D-apiofuranosyl, Rha: α-L-rhamnopyranosyl.
3
Phytomedicine 60 (2019) 152905
the activation of the TCR-MHC interaction, and the changes of the ionic
balance Na+/K+, stimulates T cell activation resulting in the secretion
of Th1 cytokines (Marciani, 2018) (Fig. 3).
Recently, studies in mouse APCs (DCs and macrophages) identified
QS-21 as an activator of the NLRP3 inflammasome. QS-21 in combi-
nation with the TLR4-agonist 3-O-deacyl-4′-monophosphoryl lipid A
(MPL A), was shown to activate the ACS-NLRP3 inflammasome, a multi-
protein complex and cause subsequent release of caspase-1 dependent
proinflammatory cytokines Il-1β/Il-18 that can promote Th 17 cell
maturation or drive INF-γ-mediated Th1 responses, respectively (Marty-
Roix et al., 2016). Thus, inflammasome signaling has the potential to
direct the development of T helper subsets. However, the role of this
signaling pathway in vivo remained to be clarified.
Structure activity relationships and semi-synthetic analogs
Although QS-21 has been used in many clinical investigations of
vaccines, several drawbacks inherent to the natural product, such as its
chemical instability, scarcity, heterogeneity, dose-limiting toxicity and
poorly understood mechanism of action have limited its widespread
clinical advancement, except for the recent malaria (Mosquirix™) and
shingles (Shingrix™) vaccines developed by GlaxoSmithKline (GSK). To
overcome these limitations, structural modifications of the natural
product through chemical synthesis have been undertaken during the
last decade, leading to the preparation of nearly 50 saponin analogs,
which have provided key insights into structure/activity relationships
(SAR) which are summarized in Fig. 4 (Fernández-Tejada et al., 2016).
These achievements allowed the identification of new novel saponins
probes with potent adjuvant activities, increased stability, and de-
creased toxicity, and the investigation into their mechanisms of action
(Fernández-Tajeda et al., 2016; Fernández-Tejada, 2017). Firstly, the
total synthesis of each isomeric QS-21 produced this potent adjuvant in
high purity and homogeneous composition. Then, some QS-21 variants
(SQS-101, SQS-102, SQS-103) were chemically synthesized with more
stable amide linkages in the acyl chain instead of the unstable native
esters (Fig. 5). Their immunological evaluation in mice demonstrated
adjuvant activities comparable to QS-21, as assessed by measuring Ab
responses by ELISA one week after booster injection (day 72) and sig-
nificantly lower toxicity than the natural product, except for SQS-102.
Further modifications in the acyl chain with a dodecanedioic acid and
truncation of the tetrasaccharide led to a trisaccharide variant with
potent adjuvant activity and whose toxicity issue is not completely
resolved (Fernández-Tejada et al., 2016). This simplification led to
further structural variations at the linker (ester linkage at C-28) be-
tween the triterpene and the linear oligosaccharide in order to
M.-A. Lacaille-Dubois
Fig. 3. Proposed mechanism of action of QS-21 (adapted from Marciani, 2018).
modulate the distance and orientation of these two domains. The var-
iants exhibit striking differences in adjuvant activity and toxicity in
mouse vaccination models, yielding new insights into the structural
requirements for adjuvant activity. Molecular dynamics simulations of
these compounds and QS-21 Api have revealed distinctive conforma-
tional features correlating with adjuvant activity, which may help in the
design of new analogs (Walkowicz et al., 2016). The modifications of
the tetrasaccharide into a trisaccharide variant, and the acyl chain into
a terminal amine variant enables the synthesis of novel acyl chain
variants bearing fluorescent and iodinated substituents for subsequent
imaging and in vivo biodistribution studies (Fernández-Tejada et al.,
2014). They were adjuvant-active in several mouse-vaccination models
and less toxic than QS-21. Early studies of biodistribution and fluores-
cence imaging allowed investigations into the enigmatic mechanism of
action of these saponins. Preliminary results in vaccinated mice sug-
gested the potential role of these fluorescent probes in Ag ovalbumin
(OVA) trafficking by APCs from the site of injection to the LNs for its
presentation to the immune system and yielded some mechanistic un-
derstanding of the potentiation of the immune response (Fernández-
Tejada et al., 2014). Extensive structure/function studies have shown
that aryl iodide derivatives with a free C-3 OH display a good efficacy/
Fig. 4. Summary of saponin structure adjuvant activity relationships (adapted from Fernández-Tejada et al., 2016).
4
Phytomedicine 60 (2019) 152905
toxicity profile. It was concluded that the trisaccharide at C-3 was not
required for adjuvant activity. Modifications at C-4 aldehyde and C-16
hydroxyl have shown that echinocystic acid aryl iodide derivatives
lacking the C-4 aldehyde but retaining the C-16 alcoholic function
(Fig. 6) exhibited potent adjuvant activity and no toxicity in a pre-
clinical vaccination model using a four-component vaccine (MUC1-
KLH, OVA, GD3-KLH) when tested at doses of 20 and 50 μg. These data
indicated that the C-4 aldehyde previously suggested to participate in
Schiff's base formation with a presumed T cell surface receptor target
(Marciani, 2003), is not required for adjuvant activity and revealed the
previously unknown role of the C-16 alcohol in enhancing activity. This
opens the door to semi-synthesis from easily available alternative pre-
cursors and allowed detailed mechanistic studies (Fernández-
Tejada et al., 2014).
QS-21 and new adjuvant formulations
Adjuvant systems
One of the promising approaches to improving efficiency of newly
developed vaccines is given by the combination of several adjuvants
M.-A. Lacaille-Dubois
Fig. 5. Structure of stable amide QS-21 structural analogs.
Fig. 6. An echinocystic acid aryl iodinated QS-21 structural analog.
into single formulations, such as the adjuvant systems AS01, AS02,
AS015 (Garçon et al., 2011). AS01, developed more than 20 years ago,
is a liposome-based adjuvant comprising two immunostimulants, MPL
and QS-21. Liposomes are synthetic nanospheres consisting of phos-
pholipid bilayers that can encapsulate antigens and act as antigen de-
livery vehicles (Garçon et al., 2011). MPL is a detoxified synthetic de-
rivative of the lipopolysaccharide (LPS) from the bacteria Salmonella
minnesota and induces activation of innate immunity via TLR-4, sti-
mulating NF-kB transcriptional activity. It stimulates the induction of
Ag-specific T cells producing interferon-γ (IFN- γ) and IgG2A anti-
bodies. QS-21 from Q. Saponaria (licensed by GSK from Antigenics Inc.,
a wholly owned subsidiary of Agenus Inc., Lexington, MA, USA) in the
early evaluations as an adjuvant was suggested to promote high Ag-
specific Ab responses and CD8+ T cell responses in mice and high Ag-
specific Ab responses in humans (Didierlaurent et al., 2014). QS-21, as
an amphiphilic compound is known for its hemolytic properties, which
can be abrogated through formulation in liposomes in the presence of
cholesterol such as in AS01 (Beck et al., 2015; Garçon and Di Pasquale,
2017). This resulted in an acceptable tolerability profile for its use in
human vaccines. AS02 contains MPL/QS-21 in an oil-in-water emul-
sion. AS015, combining QS-21, MPL, and CpG 7909 (an oligodesox-
ynucleotide and TLR-9 agonist), in a liposome is the most complex
combination of adjuvants which has been used for applications in
cancer immunotherapy.
Mechanisms of action of QS-21 when formulated in liposomes
The mechanism of action of QS-21 in a liposome is poorly under-
stood despite the clinical efficacy of AS01. Therefore, some studies were
carried out in order to tentatively elucidate the adjuvant effect of QS-21
in such a formulation. It was observed after an intramuscular (im)
immunization in mice against two model antigens, Hepatitis B surface
antigen (HBsAg) and OVA that QS-21, rapidly accumulated in CD169+
resident macrophages of the dLN, where it induces caspase-1 activation.
These early events then organized the recruitment of innate immune
5
Phytomedicine 60 (2019) 152905
cells and activation of DCs and subsequent T cell priming. Further
analysis showed that the adjuvant effect of QS-21 depended on the
integration of caspase-1 and MyD88 pathways at least in part through
local release of HMGB1 (High-Mobility-Group protein B1)
(Detienne et al., 2016).
A recent study reported that QS-21 directly activated human
monocyte-derived dendritic cells (mo-DCs) and promoted a pro-in-
flammatory transcriptional program (Welsby et al., 2017). In this
model, QS-21 was endocytosed via a cholesterol-dependent mechanism
and accumulated in lysosomes where it induces their destabilization
through pore formation and potential release of their contents. This
resulted in inflammasome activation, Syk- (a tyrosine kinase) and ca-
thepsin-B (a cysteine protease) dependent cell activation and cytokine
production in mo-DCs. Lysosomal disruption could also affect Ag pro-
cessing and translocation to the cytosol for presentation on MHCs. Fi-
nally it was shown that cathepsin B contributes to the adjuvant prop-
erties of QS-21 on both CD4+ and CD8+ Ag-specific T-cell responses in
vivo.
Mechanism of action of AS01: synergistic effect of QS-21 and MPL
The Adjuvant Systems containing MPL/QS21 in combination with
HBsAg were shown to induce very strong and persistent humoral and
cellular immune responses in healthy adults, AS01B inducing the
strongest and most durable specific CMI after two doses, without ser-
ious adverse events. The CD4+ response was characterized in vitro by
high lymphoproliferation, high IFN-γ and moderate Il-5 production.
This response was confirmed ex vivo by detection of Il-2 and IFN-γ
producing CD4+ T cells and in vivo by measuring increased levels of
IFN-γ in the serum (Vandepapelière et al., 2008). Therefore AS01B has
been selected as lead AS for the development of vaccines. Experiments
were conducted in mouse models with a recombinant varicella-zoster
virus (VZV) glycoprotein E (gE) and the fluorescently labeled QS-21
incorporated in AS01 (HZ (shingle) vaccine formulation). AS01 induces
a rapid and transient activation of the innate immune system, both at
M.-A. Lacaille-Dubois
the injection site and in the dLN, leading to the generation of high
number of efficient Ag-loaded DCs in the dLN to promote Ag-specific
adaptive immune responses (Didierlaurent et al., 2014). A comparative
study of the immunogenicity of different formulations containing the
subunit gE was carried out in a VZV-primed mouse model
(Dendouga et al., 2012). This model was chosen in order to mimic the
clinical setting, in which the large majority of persons at elevated risk
for HZ are VZV sero-positive. An HZ candidate vaccine comprising gE
(5 μg) and AS01B (5 μg each of MPL and QS-21) was shown to be highly
immunogenic in mice, inducing higher responses of CD4+ T cells ex-
pressing Il-2 and/or INF-γ and higher humoral responses compared to
other gE/AS01 formulations with different Ag doses (0.5, 2.5, 10 μg) or
with lower doses of adjuvant (2.5 and 1.25 μg each of MPL and QS-21,
corresponding to AS01E and AS01F, respectively) (Dendouga et al.,
2012). Furthermore, the contribution of each component of AS01B to
the induction of cellular and humoral response was assessed after im-
munization of mice with gE either unadjuvanted, or adjuvanted with
QS-21 (5 μg), MPL (5 μg) (both in liposomes), and AS01. The results
showed that the gE-specific cytokine (IFN-γ, Il-2) producing CD4+ T
cell responses induced by gE/liposome+QS21 and gE/liposome+MPL
were of similar and low magnitude, whereas the response induced by
gE/AS01B was significantly higher than gE/liposome+QS21 (8.7 fold
difference) and gE/liposome+MPL (7.5 fold difference). This clearly
showed that QS-21 and MPL acted synergistically to induce a high
frequency of gE-specific CD4+ T cells. For the anti-gE antibody re-
sponses, an additive effect, rather than a synergistic effect was observed
(Dendouga et al., 2012; Didierlaurent et al., 2017). These results sug-
gested that the gE/AS01B vaccine candidate was appropriate for further
clinical assessment.
An investigation of the molecular and cellular mechanism of AS01
adjuvanted vaccines was undertaken in order to clarify how im-
munostimulants function in combination. (Coccia et al., 2017). Results
from in vivo and clinical studies showed that the combination of MPL/
QS-21 in AS01 resulted in the stimulation of de novo pathways that
were not triggered by the individual components. Mice were im-
munized twice, 2 weeks apart, with the RTS,S antigen of the malaria
vaccine, formulated either without adjuvant, with MPL, with QS-21
(both included in liposomes) or with AS01. RTS,S is a recombinant
protein consisting of repeated sequences of the Plasmodium falciparum
circumsporozoite protein (CSP-repeat region, (R)) and T-cell epitope
domain (T) linked to the hepatitis B surface antigen (HBsAg) (S), and
HBsAg alone (S) (Moris et al., 2018). The CSP-specific immune response
was measured after the second dose. The results clearly showed that
MPL and QS-21 act synergistically to induce strong antibody and Ag-
specific CD4+ T cell responses. In contrast, very low levels of CD8+ T
cell responses were generated against CSP. Further investigations
highlighted the key role of the early release of IFN-γ by innate resident
cells (NK and CD8+ T cells) in the LN, draining the injection site, which
is essential for the development of the CD4+ T cell response (Th1 im-
munity). This was confirmed by measuring the levels of the cytokine
after immunization of mice with HBs adjuvanted with AS01 or with its
components. AS01 induced the early production of IFN-γ with a peak in
concentration at 6 h post immunization (300 pg/ml), whereas MPL or
QS-21 alone failed to significantly induce IFN-γ. This effect results from
the MPL/QS-21 synergy and is controlled by macrophages, Il-11 and Il-
18 (Coccia et al., 2017).
Clinical applications
QS-21 containing adjuvants have been assessed in more than 100
human clinical trials. AS01 is a key component of the recently devel-
oped malaria and HZ vaccines and other candidate vaccines under
development against infectious diseases, and cancer (Coccia et al.,
2017; Didierlaurent et al., 2017; Garçon and di Pasquale, 2017). Recent
clinical studies of prophylactic vaccines against malaria, HZ, HIV, tu-
berculosis from an efficacy, immunogenicity and safety points of view
6
Phytomedicine 60 (2019) 152905
(Table 1) and of therapeutic vaccines against melanoma, NSCLC and
Alzheimer's disease, will be reported below.
Prophylactic vaccines
Malaria
Despite effective medicines, insecticide-treated nets and indoor in-
secticide spraying, malaria killed 445,000 people in 2016, particularly
sub-Saharan African children (World Malaria Day, 2018). The Ag of the
candidate malaria vaccine RTS,S targets the CSP which is expressed on
the plasmodium sporozoite during the pre-erythrocytic stage of its life-
cycle, the stage between mosquito bite and liver infection. RTS,S
/AS02, was the first malaria vaccine candidate, in which an adjuvant
system was used (Garçon et al., 2007). However, RTS,S/AS01 was
shown in a phase II trial to be well tolerated, to induce strong humoral
and cellular immune response and to improve protection against Plas-
modium falciparum challenge in comparison to RTS,S/AS02
(Kester et al., 2009). Namely, it induced high levels of anti-CSP IgG
titers and CSP-specific polyfunctional CD4+ T cells expressing Il-2, IFN-
γ, TNFα or CD40L which have been associated with protection in the
experimental malaria challenge model in adults. Other clinical trials in
children and adults confirmed these observations and no serious ad-
verse events were reported (Agnandji et al., 2010; Olotu et al., 2011;
Ansong et al., 2011; White et al., 2013; Levast et al., 2014; Leroux-
Roels et al., 2014a). Therefore the development of RTS,S/AS02 was
stopped and RTS,S/S /AS01 was consequently selected for phase III
development (Garçon and Di-Pasquale, 2017). Analyses of data in term
of efficacy, immunogenicity and safety of Phase I-III trials including
adults, infants, and children from sub-Saharan countries were reviewed
(Agnandji et al., 2015). Clinical trials in children with newly ad-
juvanted vaccines led to the conclusion of clinical safety except for
some unexplained cases of meningitis in one phase III AS01 adjuvanted
malaria vaccine trial, which warrant further safety evaluations
(Stassijns et al., 2016). Important insights into the molecular me-
chanism of protective immunity against malaria were achieved invol-
ving additional immunogenicity analyses into the potential contributive
roles of CSP-specific Abs and CSP-specific CMI to protection
(Moris et al., 2018; Kazmin et al., 2017) leading to the identification of
the NF-kB and IFN-γ pathways (van den Berg et al., 2017).
The first clinical large-scale Phase III trial (The RTS,S clinical trials
partnership, 2014) evaluating a malaria vaccine involving 15,460 in-
fants and young children was completed in December 2013 at 11 sites
from 7 African countries (Burkina Faso, Gabon, Ghana, Kenya, Malawi,
Mozambique, and Tanzania). Safety and efficacy studies of the RTS,S/
AS01 vaccine (Mosquirix™) showed that it prevented many cases of
clinical and severe malaria over the 18 months after dose 3. Vaccine
efficacy was higher in children (5–17 months) than in infants (6–12
weeks), but even at modest levels (45.7%). The final results in 2015
(The RTS,S clinical trials partnership, 2015) from the same trial,
showed the enhanced efficacy by administration of a booster dose in
both age categories, 18 months after the first three injections. With
regards to risks, the safety of Mosquirix is similar to that of other
vaccines, the most common side effects being fever, pain and swelling
at the injection sites. Thus, the vaccine has the potential to make a
substantial contribution to malaria control when used in combination
with other effective control measures, especially in areas of high
transmission. The ratio benefit/risk of Mosquirix was found to be ac-
ceptable by the European Medicines Agency's committee for Medicinal
Products for Human use (EMA/CHMP) which delivered a positive sci-
entific opinion in 2015 for use outside the European Union (EU) (EMA
Press release, 2015). Furthermore, the World Health Organization re-
commended a large-scale pilot implementation of the vaccine in young
children with a four-dose schedule in African regions of moderate to
high parasite transmission (WHO, 2016). Three countries, Ghana,
Kenya, and Malawi were selected by the WHO in 2017 for this large-
scale pilot program which started in 2018. It will be the first malaria
 Table 1
Clinical trials involving newly adjuvanted prophylactic vacccines.
Phase Year Country Study populations Nr of Summary Reference
subjects
M.-A. Lacaille-Dubois

Malaria
IIA 2003–06 USA Healthy volonteers 102 RTS,S/AS01 (50% efficacy) well tolerated, safe, induced stronger Kester et al., 2009
humoral and cellular immune response, and improved protection over
RTS,S/AS02 (32% efficacy). Both vaccines were well tolerated and safe.
II 2007 Ghent, Belgium Healthy naïve adults 36 Stronger immune response of RTS,S/AS01 as compared to RTS,S/AS02, Leroux et al., 2014a
acceptable safety profile.
II 2006–08 Ghana 5–17 months old children 540 RTS,S/AS01 induced higher CD4+ T cells responses as compared to Ansong et al., 2011
RTS,S/AS02, when given on a 1,2, 7-months schedule.
IIB 2007–08 Kenya, Korogwe,Tanzania, Kilifi 5–17 months old children 894 RTS,SAS01E provides sustained protection from clinical malaria for at Olotu et al., 2011
least 15 months. The frequency of RTS,S-induced CSP-specific TNF-α+
CD4+ T cells and the anti-CSP Ab responses were associated with
protection.
II 2007–09 Ghana, Gabon, Tanzania 6–10 weeks infants 511 RTS,S/AS01E integrated in the EPI programme of immunization showed Agnandji et al., 2010
a favorable safety and immunogenicity evaluation.
III 2009–14 Multicentric (11 sites in Africa) 6 weeks to 17 month infants 15,460 RTS,S/AS01: 3 doses at month 0, 1, 2. 18 months following vaccination: RTS,S clinical trial partnership
and children substantial contribution to. malaria control. 2014, 2015
Efficacy was enhanced by administration of a booster dose at month 20:
the vaccine prevented a substantial number of cases of clinical malaria
over a 3–4 year period.
Herpes Zoster
I/II 2004–05 Ghent Adults 18–30 years 20 gE/AS01B: highly immunogenic with CD4+ T cell and humoral immune Leroux et al., 2012
responses being more immunogenic than a live attenuated vaccine;
clinically acceptable safety profile.
Extension 2007–08 Adults 50–70 years 135

7
II 2007–10 Czech Republic, Germany, Sweden, Adults>60 years 715 gE/AS01B: three formulations containing 25 μg, 50 μg, or 100 μg of the Chlibek et al., 2014
Netherlands adjuvanted vaccine were immunogenic and well tolerated in older
adults. This vaccine induced robust CD4+ T cell and humoral immune
responses that persisted for up to 3 years after vaccination.
II follow-up study 2011–13 Czech Republic, Germany, Sweden, Adults>60 years 129 In this long term follow up study, gE-specific humoral & cellular immune Chlibek et al., 2016
Netherlands responses persisted for 6 years after two doses (2 months apart) of the
50 μg gE/AS01B formulation.
III 2010–15 18 countries North America, Europe, Adults > 50 years 15,411 High efficacy (97.2%) in preventing HZ and its complications in Lal et al., 2015
Latin America, Asia-pacific adults > 50 years, with an acceptable safety profile. Robust immune
responses persisting for 3 years following vaccination.
> 70 years 13,900 Hz/su reduces the risk of HZ (91.3%) and PHN (88.8%)among adults > Cunningham et al., 2016
70 years of age.
II (long term follow-up 2016–18 Germany, Sweden, Czech Republic Adults >60 years 70 HZ/su induces cell mediated and humoral immunity persisting up to 9 Schwarz et al., 2018
extension study) years post-initial vaccination, which were similar to those observed after
6 years. Immune responses are predicted to remain up to 15 years post
initial vaccination.
Tuberculosis
I/II 2006–07 Ghent, Belgium Mtb naïve adults 110 M72/AS01 and MF72/AS02 TB vaccine formulations: clinically well Leroux-Roels et al., 2013
tolerated and induced high magnitude of polyfunctional MF-72-specific
CD4+ T cell and Ab responses persisting at 3 years, with the highest
CD4+ T cell response for M72/AS01.
II 2010–12 Gambia BCG-vaccinated infants 300 M72/AS01: concomitantly with or after the Expanded Programme of Ikodo et al., 2014
Immunization vaccines.
acceptable safety profile; higher M72-specific humoral and CD4+ T cell
responses after 2 doses.
II 2009–10 TB endemic region Healthy, HIV-neg. 60 M72/AS01E: clinically acceptable safety and immunogenicity profile. Penn-Nicholson et al., 2015
adolescents Robust T cell (CD4+ and CD8 + T cell responses) and Ab responses.
Good candidate for advancement into efficacy trials
(continued on next page)
Phytomedicine 60 (2019) 152905
M.-A. Lacaille-Dubois Phytomedicine 60 (2019) 152905

vaccine provided to young children through routine immunization

programmes (Malaria vaccine implementation programme, 2019). The
Van Der Meeren et al., 2018
results of two phases (2017–20, and 2020–21) providing insights on the

Van Braeckel et al., 2011

Leroux Roels et al., 2010
feasibility of delivering the vaccine in real-life settings and on its safety

Leroux et al., 2014b

profile in the context of routine use, will contribute to the future de-
cisions on the wider-scale use of the vaccine.
Reference

Herpes zoster
Herpes zoster (HZ), also known as shingles is a common, painful
disease occurring principally in adults aged > 50 years and im-
munocompromised individuals. HZ is caused by the reactivation of the
of study vaccine) elicited strong, persistent and broadly reactive CD4+ T

immune responses, but no significant CD8+ T cell responses. Clinically

Gp120/Nef/Tat protein Ags adjuvanted with AS02 or AS01 (four doses

chloroquine induced a strong humoral and polyfunctional CD4+ T cell

F4/AS01: high and long-lasting frequencies of polyfunctional CD4+ T

latent varicella-zoster virus (VZV) in the dorsal root or cranial ganglia

cell responses which were more pronounced with AS01. Acceptable
M72/AS01E provided 54% protection against active pulmonary TB

usually many years after a primary VZV infection (chickenpox) that

F4/AS01B: A booster dose administered alone or two days after

occurs during childhood (Chlibek et al., 2016). HZ can lead to serious

cell responses characterized by CD40L, Il-2, INF-α, INF-γ.

complications including post herpetic neuralgia (PHN). A phase I/II

trial showed that a subunit vaccine containing the recombinant VZV
glycoprotein E (gE) as Ag and AS01B (called HZ/su) was well tolerated
and highly immunogenic with CD4+ T cell and humoral immune re-
sponses, being more immunogenic than a live attenuated vaccine
(Leroux-Roels et al., 2012). A phase II trial in adults > 60 years of age
evaluating different formulations (containing 25, 50, or 100 μg gE
without evident safety concerns

combined with AS01) and different schedules showed that all for-
mulations elicited robust humoral and CD4+ T cell immune responses
acceptable safety profile

that persisted for at least 3 years after vaccination. Two doses of gE/
AS01B were more immunogenic than one (Chlibek et al., 2014). In a
follow-up study conducted on 129 healthy adults, it was shown that the
safety profile.

gE specific CD4+ T cell and anti-gE antibody responses persisted for 6

years after two doses of 50 μg gE/AS01B, without any safety concerns
Summary

(Chlibek et al., 2016). Therefore, the dose of 50 μg gE was selected for

other clinical developments. A two randomized placebo-controlled
phase III efficacy trials in 18 countries evaluated the efficacy and safety
of HZ/su as compared to placebo in older adults (>50 and > 70 years
subjects

of age). Participants received two im doses of the vaccine or placebo, 2

Nr of

3595

180

180

28

months apart. HZ/su has demonstrated high efficacy (97.2% and 91.3%
for adults > 50 years and > 70 years, respectively) in preventing HZ
Healthy seronegative adults
HIV-negative young adults

and PHN in all age groups with an acceptable safety profile (Lal et al.,
with latent Mtb infection

HIV-seronegative adults

2015; Cunningham et al., 2016). The robust immune responses re-

Young healthy adults

mained for 3 years after vaccination as assessed in subsets of partici-

Study populations

pants of the phase III trials. Persistent anti-gE Ab and gE-specific

polyfunctional CD4+ T cell responses are likely important mechanisms
by which HZ/su confers the high efficacy against HZ
(Cunningham et al., 2018). In an extension study of the original phase II
trials reported by Chlibek et al. (2014, 2016), it was observed that HZ/
su induces cell mediated and humoral immunity persisting up to 9 years
post initial vaccination in adults > 60 years old, confirming statistical
prediction models based on immune responses measured at earlier time
Kenya, south Africa, Zambia

points. Immune responses are predicted to persist up to 15 years after

Single center in Belgium

the initial vaccination (Schwarz et al., 2018). The acceptable benefit/

risks profile led to the licensing by the U.S. Food and Drug Adminis-
tration (FDA) in 2017 of the HZ/su vaccine (Shingrix™) and a marketing
Ghent, Belgium

Ghent, Belgium

authorization valid throughout the EU on 21 March 2018 (Shingrix,

INN-herpes zoster vaccine. EMA/76073/2018). Shingrix is preferred to
Country

Zostavax (a live attenuated vaccine), providing strong protection

against shingles and PHN.
2014–2018

2007–08

2007–08

2009–10

Tuberculosis
First introduced in 1921, the Bacillus Calmette-Guérin (BCG) vac-
Year

cine is the only currently licensed vaccine available to protect against

tuberculosis (TB), a disease caused by the bacteria Mycobacterium tu-
Randomized double-blind

berculosis (M.tb.). It protects children from meningeal and severe forms

Table 1 (continued)

disseminated of TB and death, but offers limited protection against

pulmonary TB in adolescents and adults, which is the form of the dis-
ease responsible for the vast majority of transmission and TB-related
study

morbidity and mortality. Therefore, there is an urgent need for a new

Phase

and improved vaccine against TB for controlling this disease that con-
HIV

I/II
IIb

II

tinues to pose a global health threat with 1.7 million deaths in 2016

8
M.-A. Lacaille-Dubois
(Van Der Meeren et al., 2018). The M72/AS01 candidate vaccine con-
tains the recombinant fusion protein derived from the two im-
munogenic M. tuberculosis Ags (Mtb32A and Mtb39A) in AS01. A first
phase I/II randomized, controlled clinical study with M72/AS01 was
carried out in Belgium in healthy adults volunteers. The results showed
that the vaccine was clinically well tolerated and induced high and
persistent (up to three years) specific Th1 CD4+ T-cell (co-expressing
CD40L, Il-2, TNF-α and IFN-γ) and Ab responses, supporting its further
clinical evaluation in TB-endemic settings (Leroux-Roels et al., 2013).
Similar conclusions were drawn in a phase II controlled trial conducted
on healthy, HIV uninfected adolescents living in an area with high TB
burden in South Africa (Penn-Nicholson et al., 2015). In a phase II study
M72/AS01 given to infants after or concomitantly with expanded-
Programme-on-Immunization (EPI) vaccines had an acceptable safety
profile. These results suggested no interference of immunogenicity
profiles occurred following co- administration of M72/AS01 and EPI
vaccines. Two M72/AS01 doses elicited higher immune responses than
one dose (Ikodo et al., 2014). In a large phase IIb trial conducted on
3600 HIV-uninfected adults latently infected with Mtb in three African
countries, M72/AS01E was shown to provide 54% protection for Mtb-
infected adults against active pulmonary TB, without evident safety
concerns (van der Meeren et al., 2018). These results suggest further
evaluations of M72/AS01 as a possible vaccination strategy against TB.
HIV-infections
There were approximately 36.9 million people living with HIV at
the end of 2017 with 1.8 million people becoming newly infected in
2017 (HIV/AIDS, Key facts, 19 July 2018). While treatments are
available to treat or to prevent HIV infections, there is no vaccine
currently licensed. Despite ongoing prevention efforts, there is an ur-
gent need for a safe and effective prophylactic vaccine. Prevention of
infection through induction of neutralizing Abs, induction of strong
cellular immune responses in order to delay progression and reduce the
transmission rate in high risk population, are the major aims for an HIV
vaccine. Although clinical attempts were disappointing, they con-
tributed valuable insights into immune protective immunity.
A randomized double-blind study conducted on healthy HIV-ser-
onegative adults showed that a vaccine formulation containing gp 120,
Nef and Tat Ags and AS01B induced strong Ab response and CD4+ T cell
responses characterized by high lymphoproliferative capacity and Il-2
production, that were maintained up to 18 months after the last vaccine
dose. A CD8+ T cell response was not observed (Leroux-Roels et al.,
2010). This study conducted with different adjuvants underlined the
superiority of responses with AS01B in comparison with other ad-
juvants, confirming previous findings. Another candidate HIV-1 vaccine
consisting of a recombinant fusion protein (F4) encoding four HIV-1
clade B Ags (p17, p24, reverse transcriptase (RT) and Nef) adjuvanted
with AS01 was studied in a phase I/II trials with healthy HIV-ser-
onegative volunteers (Van Braeckel et al., 2011). This vaccine was
shown to be immunogenic with an acceptable safety profile. After two
doses of 10 μg, strong polyfunctional (CD40L, Il-2, TNF-α, IFN-γ phe-
notype) and persistent CD4+ T cell responses were induced, suggesting
that this vaccine merits further evaluation both in a prophylactic setting
and as a potentially disease-modifying therapeutic vaccine in HIV-1-
infected subjects. A phase I clinical study in such subjects has been
initiated (Harrer et al., 2014). A phase II study was conducted with the
F4/AS01B candidate vaccine on healthy adults in order to evaluate the
effect of chloroquine on the specific CD8+ T cell responses and the
safety profile of a booster dose (Leroux-Roels et al., 2014b). The results
showed that a booster dose, administered alone or two days after
chloroquine induced a strong Ab immune response and robust F4
-specific CD4+ T cell response, but no significant CD8+ T cell response.
This suggest that two primary doses of the F4/AS01B vaccine induce
long-lasting memory B cell and CD4+ T cell responses in healthy adults,
confirming previous findings of the phase I/II study (Leroux-
Roels et al., 2014b). This vaccine was shown to be immunogenic with a
9
Phytomedicine 60 (2019) 152905
clinically acceptable safety profile, but doesn't show significant viral
efficacy in antiretroviral therapy-naïve HIV-1-infected adults, results
confirmed in a long term study (Dinges et al., 2016; Harrer et al., 2018).
Therapeutic vaccines
Cancer
The use of immunotherapeutic vaccines in cancer patients is aimed
at inducing immune responses against existing tumors rather than
protecting healthy individuals. They have been used in phase II and III
trials against melanoma and lung cancer after surgical resection, and
the disappointing results of the most recent studies will be shortly
presented here. Enhanced production of ganglioside GM2 is observed in
different human tumor types including melanoma. Anti-GM2 vaccines
such as GM2 conjugated to a carrier keyhole limpet hemocyanin (KLH)
and administered with QS-21 induced high IgM and IgG Ab responses
but failed to show a beneficial effect in melanoma patients as it was
shown in a phase III clinical trial in term of relapse free survival, distant
metastasis-free survival and overall survival (Eggermont et al., 2013). A
five-years survival occurs for less than 50% of the patients with com-
pletely resected non-small-cell-lung cancer (NSCLC). Therapeutic
cancer vaccination has the potential for eliminating remaining cancer
cells after complete resection, thereby decreasing relapse rates and
improving survival (Vansteenkiste et al., 2016). Ag-specific im-
munotherapies enhance T-cell responses against specifically expressed
tumor Ags. In this context, the use of MAGE-A3 (Melanoma AntiGEn
family A, 3) as Ag, which is expressed in a wide array of malignancies
can be an application in cancer immunotherapy (Esfandiary and
Ghafouri-Fard, 2015). The MAGE-A3/AS015 immunotheraputic vac-
cine was assessed for clinical efficacy in an international, multicenter
phase III study in surgically resected NSCLCs. This study confirmed the
acceptable clinical safety profile of the vaccine, but did not improve
overall survival. Therefore, its further development for use in NSLCs has
been stopped (Vansteenkiste et al., 2016). Similar conclusions were
drawn from a phase III trial of a MAGE-A3/AS015 vaccine in patients
with resected, MAGE-A3-positive, stage III melanoma, which led to the
arrest of the clinical development of this vaccine for use in melanoma
(Dreno et al., 2018).
Alzheimer's disease
The pathogenesis of Alzheimer disease (AD) is associated with the
accumulation of amyloid β (Aβ) and/or hyperphosphorylated tau pro-
teins in the brain. Active immunotherapy of AD is the most extensively
studied approach, in order to evaluate its ability to elicite anti-Aβ an-
tibodies, to induce Aβ clearance, and reduce Aβ accumulation in the
brain. Vaccination with a full length Aβ peptide (Aβ1-42) successfully
elicited anti-Aβ-Ab in human subjects with AD, but adverse effects such
as meningoencephalitis were observed, attributed to T cell activation
(Winblad et al., 2014). To avoid this safety issue, a N-terminal (Aβ1−7)
peptide conjugated to a carrier protein called vanutide cridificar (ACC-
001) was produced. In preclinical studies, it was shown to generate N-
terminal Aβ Abs in adult nonhuman primates without inducing Aβ-di-
rected T cell response, supporting further clinical studies. Several phase
II trials in patients with mild to moderate AD were conducted in USA,
Europe, and Japan by using 3 doses of ACC-001 (3, 10 and 30 μg) with
and without the adjuvant QS-21 (50 μg) or placebo to investigate dose-
range, safety, immunogenicity and long term treatment (Arai et al.,
2015; Pasquier et al., 2016). ACC-001+QS-21 elicited high, sustained
anti-Aβ IgG Ab titers compared with ACC-001 alone, highlighting the
role of QS-21 in this effect. No difference was observed among the three
doses tested. Furthermore, ACC-001 at all dose levels with or without
QS-21 was generally safe and well tolerated, but exploratory cognitive
evaluations did not show differences between treatment groups and
placebo. Similar safety, tolerability and anti-Aβ- IgG immunogenicity
profiles were observed in long term phase II extensions studies, sug-
gesting that side effects do not limit in principle anti-amyloid active
M.-A. Lacaille-Dubois
immunotherapy (Hüll et al., 2017). However, larger trials of adequate
duration, with optimized dosing may be needed to attest efficacy of
anti-Aβ vaccine therapy for AD.
Conclusion
This overview provides a summary of recent reports on the me-
chanism of action of QS-21, a promising triterpene glycoside vaccine
adjuvant isolated from Quillaja Saponaria and the most relevant clinical
applications. This compound showed a potent adjuvant activity sti-
mulating humoral and cell-mediated immunity against a wide variety of
Ags and presented advantages over aluminium salts by inducing a Th1-
type immune response, necessary for controlling most intracellular
pathogens. It is unlikely that a single mechanism of action could explain
its effects as an adjuvant.
This contribution highlights:
1) the role of QS-21 acting on both APCs and T cells. The interaction
between the aldehyde function and T-cell receptors such as CD2,
delivering co-stimulatory signals together with the presentation of
the Ag peptides by APCs to the T-cell resulted in the activation of T
lymphocytes and secretions of Th1 cytokines, an important im-
mune effector mechanism for the elimination of infected cells.
2) a novel mechanism of action based on the activation of caspase 1-
dependent NLRP3 inflammasome in mouse APCs, leading to the
secretion of proinflammatory cytokine Il-1β/Il-18, which are im-
portant for Th1 responses.
3) the synthesis of nearly 50 structural analogs of QS-21 and detailed
SAR studies leading to more easily available, and stable molecules
with an improved activity/toxicity profile, among them a novel
aryl iodinated simplified echinocystic derivative showing a potent
adjuvant activity and considerably attenuated toxicity.
4) the signaling pathways of QS-21 when formulated in a liposome,
such as the rapid accumulation in dLN resident CD169+ macro-
phages, where it induces caspase-1 activation and HMGB1 release.
These events orchestrate the recruitment of innate immune cells
and activation of DCs leading to T cell priming. A direct activation
of DCs was also reported, resulting in release of cathepsin B which
contribute to specific T cell responses in vivo.
5) the development of Adjuvant Systems (AS) including a combina-
tion of immunostimulants in a formulation, such as AS01 (QS-21/
MPL in a liposome), in which they were shown to act synergisti-
cally in enhancing CMI responses.
6) the high number of clinical applications in term of im-
munogenicity, efficacy, safety of QS-21 alone or in adjuvant sys-
tems in many prophylactic vaccines against complex pathogens,
e.g. malaria, herpes zoster, tuberculosis, HIV, and therapeutic
vaccines, e.g. cancer, AD.
7) the licensing by the US FDA in 2017 of a vaccine against shingles
called HZ/su (Shingrix™) containing gE/AS01 and its commercia-
lization in the EU since march 2018.
8) the first malaria vaccine (Mosquirix™) containing RTS,S/AS01 re-
ceiving a regulatory approval by the EMA in 2015 for use outside of
Europe and being currently under exploration in sub-Saharan re-
gions for its use in routine.
9) the disappointing phase III clinical results in term of disease-free
survival of patient with immunotherapeutic vaccines adjuvanted
with QS-21/AS015 against NSCLC and melanoma despite pro-
mising results of phase II trials.
10) the encouraging results of a phase II trial confirming that the long
term treatment with the immunotherapeutic vaccine vanutide cri-
dificar (ACC-001) adjuvanted with QS-21 against Alzheimer's dis-
ease was well tolerated with an acceptable safety profile.
10
Phytomedicine 60 (2019) 152905
Perspectives
The development of adjuvants in vaccines represents an area in
constant evolution with substantial advances over the past two decades.
QS-21 in ASs is a key component of multiple vaccines targeting infec-
tions and endemic diseases in developing countries. Recent research has
led to the commercialization of Shingrix™ and the approval of
Mosquirix™, two vaccines against HZ in adults and malaria in children,
respectively, which contain QS-21 as a part of their formulation. To
ensure a continuous supply of this exciting molecule, the researchers
aimed in the near future to find an alternative innovative process of
producing this compound on a large scale based on plant cell-culture.
This might overcome some limitations of the current traditional tech-
niques of extraction with tedious purification processes and chemical
synthesis involving many complex reactions. However, the synthetic
technology initially used for the first synthesis of QS-21, was subse-
quently applied to preparing novel structural analogs with potent ac-
tivity and low toxicity, leading to SAR studies and a better under-
standing of the mode of action. Further work in this area could facilitate
novel saponin design that will lead to the discovery of new adjuvants
and improved adjuvant-antigen combinations for future vaccines, par-
ticularly against infectious tropical diseases such as Dengue,
Leishmaniasis, and Chagas diseases, which continue to cause significant
morbidity and mortality in developing countries.
Conflict of interest
There are no conflicts of interest associated with this publication.
References
Agnandji, S.T., Asante, K.P., Lyimo, J., Vekemans, J., Soulanoudjingar, S.S., Owusu, R.,
Shomari, M., Leach, A., Fernandes, J., Dosoo, D., et al., 2010. Evaluation of the safety
and immunogenicity of the RTS,S/AS01E malaria candidate vaccine when integrated
in the expanded program of immunization. J. Infect. Dis. 202, 1076–1087.
Agnandji, S.T., Fernandez, J.F., Bache, E.B., Ramharter, M., 2015. Clinical development
of RTS,S/AS malaria vaccine, a systematic review of clinical phase I-III trials. Future
Microbiol. 10, 1553–1578.
Ansong, D., Asante, K.P., Vekemans, J., Owusu, S.K., Owusu, R., Brobby, N.A.W., Dosoo,
D., Osei-Akoto, A., Osei-Kwakye, K., Asafo-Adjei, E., et al., 2011. T cell responses to
the RTS,S/AS01E and RTS,S/AS02D malaria candidate vaccines administered ac-
cording to different schedules to Ghanaian children. PLoS One 6, e18891.
Arai, H., Suzuki, H., Yoshiyama, T., 2015. Vanutide cridificar and the QS-21 adjuvant in
Japanese subjects with mild to moderate Alzheimer disease: results from two phase 2
studies. Curr. Alzheimer Res. 12, 242–254.
Awate, S., Babiuk, L.A., Mutwiri, G., 2013. Mechanisms of action of adjuvants. Front.
Immunol. 4 (114), 1–10.
Beck, Z., Matyas, G.R., Alving, C.R., 2015. Detection of liposomal cholesterol and
monophosphoryl lipid A by QS-21 saponin and Limulus polyphemus amebocyte lysate.
Biochim. Biophys. Acta 1848, 775–780.
Chlibek, R., Smetana, J., Pauksens, K., Rombo, L., Van den Hoek, J.A., Richardus, J.H.,
et al., 2014. Safety and immunogenicity of three different formulations of an ad-
juvanted varicella-zoster virus subunit vaccine in older adults: a phase II, randomized
controlled study. Vaccine 32, 1745–1753.
Chlibek, R., Pauksens, K., Rombo, L., van Rijckevorsel, G., Richardus, J.H., Plassmann, G.,
Schwarz, T.F., Catteau, G., Lak, H., Heineman, T.C., 2016. Long-term im-
munogenicity and safety of an investigational herpes zoster subunit vaccine in older
adults. Vaccine 34, 863–868.
Coccia, M., Collignon, C., Hervé, C., Chalon, A., Welsby, I., Detienne, S., van Helden, M.J.,
Dutta, S., Genito, C.J., Waters, N.C., et al., 2017. Cellular and molecular synergy in
AS01-adjuvanted vaccines results in an early IFNγ response promoting vaccine im-
munogenicity. NPJ Vaccines 2, 25.
Cunningham, A.L., Lal, H., Kovac, M., Chlibek, R., Hwang, S.J., Diez-Domingo, J.,
Godeaux, O., Levin, M.J., McElhaney, J.E., Puig-Barbera, J., et al., 2016. Efficacy of
Herpes–Zoster subunit vaccine in adults 70 years of age or older. N. Engl. J. Med.
375, 1019–1032.
Cunningham, A.L., Heineman, T.C., Lal, O., Godeaux, O., Chlibek, R., Hwang, S.J.,
McElhaney, J.E., Vesikari, T., Andrews, C., Choi, W.S., et al., 2018. Immune responses
to a recombinant glycoprotein E Herpes Zoster vaccine in adults aged 50 years or
older. J. Infect. Dis. 217, 1750–1760.
Del Giudice, G., Rappuoli, R., Didierlaurent, A.M., 2018. Correlates of adjuvanticity: a
review on adjuvants in licenced vaccines. Semin. Immunol. 39, 14–21.
Dendouga, N., Fochesato, M., Lockman, L., Mossman, S., Giannini, S.L., 2012. Cell-
mediated immune responses to a varicella-zoster virus glycoprotein E vaccine using
both a TLR agonist and QS-21 in mice. Vaccine 30 3126–3135.
Detienne, S., Welsby, I., Collignon, C., Wouters, S., Coccia, M., Delhaye, S., Van Maele, L.,
M.-A. Lacaille-Dubois
Thomas, S., Swertvaegher, M., Detavernier, A., et al., 2016. Central role of CD169+
lymph node resident macrophages in the adjuvanticity of the QS-21 component of
AS01. Sci. Rep. 6 (39475), 1–14.
Didierlaurent, A.M., Collignon, C., Bourguignon, P., Wouters, S., Fierens, K., Fochesato,
M., Dendouga, N., Langlet, C., Malissen, B., Lambrecht, B.N., et al., 2014.
Enhancement of adaptive immunity by the human vaccine adjuvant AS01 depends on
activated dendritic cells. J. Immunol. 193, 1920–1930.
Didierlaurent, A.M., Laupèze, B., Di Pasquale, A., Hergli, N., Collignon, C., Garçon, N.,
2017. Adjuvant system AS01: helping to overcome the challenges of modern vac-
cines. Expert Rev. Vaccines 16, 55–63.
Dinges, W., Girard, P.M., Podzamczer, D., Brockmeyer, N.H., Garcia, P., Harrer, T.,
Lelievre, J.D., Franck, I., Colin de Verdière, N., Yeni, G.P., et al., 2016. The F4/AS01B
HIV-1 vaccine candidate is safe and immunogenic, but doesn't show viral efficacy in
antiretroviral therapy-naïve, HIV-1 infected adults: a randomized controlled trial.
Medicine 95 (6), e2673.
Di Pasquale, A., Preiss, S., Tavares Da Silva, F., Garçon, N., 2015. Vaccine adjuvants: from
1920 to 2015 and beyond. Vaccines 3, 320–343.
Dreno, B., Thompson, J.F., Smithers, B.M., Santinami, M., Jouary, T., Gutzmer, R.,
Levchenko, E., Rutkowski, P., Grob, J.J., Korovin, S., et al., 2018. MAGE-A3 im-
munotherapeutic as ajuvant therapy for patients with resected, MAGE-A3-positive,
stage III melanoma (DERMA): a double blind, randomized, placebo-controlled, phase
3 trial. Lancet Oncol. 19, 916–929.
Eggermont, A.M.M., Suciu, S., Rutkowski, P., Marsden, J., Santinami, M., Corrie, P.,
Aamdal, S., Ascierto, P.A., Patel, P.M., Kruit, W.H., et al., 2013. Adjuvant ganglioside
GM2-KLH/QS-21 vaccination versus observation after resection of primary tumor
>1.5 mm in patients with stage II melanoma: results of the EORTC 18961 rando-
mized phase III trial. J. Clin. Oncol. 31, 3831–3837.
Esfandiary, A., Ghafouri-Fard, S., 2015. MAGE-A3: an immunogenic target used in clinical
practice. Future Med. 7, 683–704.
Fernández-Tejada, A., 2017. Design, synthesis and evaluation of optimized saponin var-
iants derived from the vaccine adjuvant QS-21. Pure Appl. Chem. 89, 1359–1378.
Fernández-Tejada, A., Chea, E.K., George, C., Pillarsetty, N., Gardner, J.R., Livingston,
P.O., Ragupathi, G., Lewis, J.S., Tan, D.S., Gin, D.Y., 2014. Development of a minimal
saponin vaccine adjuvant bsed on QS-21. Nat. Chem. 6, 635–643.
Fernández-Tejada, A., Tan, D.S., Gin, D.Y., 2016. Development of improved vaccine ad-
juvants based on the saponin natural product QS-21 through chemical synthesis. Acc.
Chem. Res. 49, 1741–1756.
Garçon, N., Di Pasquale, A., 2017. From discovery to licensure, the adjuvant system story.
Hum. Vaccin. Immunother. 13, 19–33.
Garçon, N., Chomez, P., Van Mechelen, M., 2007. GlaxoSmithKline Adjuvant Systems in
vaccines: concepts, achievements and perspectives. Expert Rev. Vaccines 6, 723–739.
Garçon, N., Leroux-Roels, G., Cheng, W.F., 2011. Vaccine adjuvants. Understanding
Modern Vaccines: Perspectives in Vaccinology 1, 89–113.
Guy, B., 2007. The perfect mix: recent progress in adjuvant research. Nat. Rev. Microbiol.
5, 505–517.
Harrer, T., Plettenberg, A., Arasteh, K., Van Lunzen, J., Fätkenheuer, G., Jaeger, H.,
Janssens, M., Burny, W., Collard, F., Roman, F., et al., 2014. Safetey and im-
munogenicity of an adjuvanted protein therapeutic HIV-1 vaccine in subjects with
HIV-1 infection: a randomized placebo-controlled study. Vaccine 32, 2657–2665.
Harrer, T., Dinges, W., Roman, F., on behalf of TH-HIV-011 study group, 2018. Long-term
follow-up of HIV-1-infected adults who received the F4/AS01B HIV-1 vaccine can-
didate in two randomized controlled trials. Vaccine 36, 2683–2686.
Hüll, M., Sadowsky, C., Arai, H., Le Prince Leterme, G., Holstein, A., Booth, K., Peng, Y.,
Yoshiyama, T., Suzuki, H., Ketter, N., et al., 2017. Long term extensions of rando-
mized vaccination trials of ACC-001 and QS-21 in mild to moderate Alzheimer's
disease. Curr. Alzheimer Res. 14, 696–708.
Ikodo, O.T., Owolabi, O.A., Owiafe, P.K., Moris, P., Odutola, A., Bollaerts, A., Ogundare,
E., Jongert, E., Demoitié, M.A., Ofori-Anyinam, O., et al., 2014. Safety and im-
munogenicity of the M72/AS01 candidate tuberculosis vaccine when given as a
booster to BCG in Gambian infants: an open-label randomized controlled trial.
Tuberculosis 94, 564–578.
Kazmin, D., Nakaya, H.I., Lee, E.K., Johnson, M.J., van der Most, R., van der Berg, R.,
Ripley Ballou, W., Jongert, E., Wille-Reece, U., Ockenhouse, C., et al., 2017. System
analysis of protective immune responses to RTS,S malaria vaccination in humans.
PNAS 114, 2425–2430.
Kensil, C.R., 1996. Saponins as vaccine adjuvants. Crit. Rev. Ther. Drug Carrier Syst. 13,
1–55.
Kensil, C.R., Patel, U., Lennick, M., Marciani, D., 1991. Separation and characterization of
saponins with adjuvant activity from Quillaja saponaria Molina cortex. J. Immunol.
146, 431–437.
Kester, K.E., Cummings, J.F., Ofori-Anyinam, O., Ockenhouse, C.F., Krzych, U., Moris, P.,
Schwenk, R., Nielsen, R.A., Debebe, Z., Pinelis, E., et al., 2009. Randomized, double-
blind, phase 2a trial of falciparum malaria vaccines RTS,S/AS01B and RTS,S/AS02A
in malaria-naïve adults: safety, efficacy, and immunologic associates of protection. J.
Inf. Dis. 2009, 337–346.
Lacaille-Dubois, M.-A., Wagner, H., 1996. A review of the biological and pharmacological
activities of saponins. Phytomedicine 2, 363–386.
Lacaille-Dubois, M.-A., Wagner, H., 2000. Bioactive saponins from plants: an update. In:
Atta-Ur-Rahman (Ed.), Studies in Natural Products Chemistry Series 21. Elsevier
Science, pp. 633–687.
Lacaille-Dubois, M.-A., Wagner, H., 2017. New perspectives for natural triterpene gly-
cosides as potential adjuvants. Phytomedicine 37, 49–57.
Lacaille-Dubois, M.A., 1999. Saponins as immunoadjuvants and immunostimulants. In:
Wagner, H. (Ed.), Immunomodulatory Agents from Plants. Birkhäuser Verlag, Basel,
Switzerland, pp. 243–272.
Lacaille-Dubois, M.A., 2005. Bioactive saponins with cancer related and
11
Phytomedicine 60 (2019) 152905
immunomodulatory activity: recent developments. Stud. Nat. Prod. Chem. 32,
209–246.
Lal, H., Cunningham, A.L., Godeaux, O., Chlibek, R., Diez-Domingo, J., Hwang, S.J.,
Levin, M.J., Mc ElHaney, J.E., Poder, A., Puig-Barbera, J., Vesikari, T., Daisuke, D.,
et al., 2015. Efficacy of an adjuvanted Herpes Zoster subunit vaccine in older adults.
N. Engl. J. Med. 372, 2087–2096.
Lee, S., Nguyen, M.T., 2015. Recent advances of vaccine adjuvants for infectious diseases.
Immune Netw. 15, 51–57.
Leroux-Roels, G., Leroux-Roels, I., Clément, F., Ofori-Anyinam, O., Lievens, M., Jongert,
E., Moris, P., Ripley Ballou, W., Cohen, J., 2014Aa. Evaluation of the immune re-
sponse to RTS,S/AS01 and RTS,S/AS02 adjuvanted vaccines. Hum. Vaccines
Immunother. 10, 2211–2219.
Leroux-Roels, G., Bourguignon, P., Willekens, J., Janssens, M., Clement, F., Didierlaurent,
A.M., Fissette, L., Roman, F., Boutriau, D., 2014Ab. Immunogenicity and safety of a
booster dose of an investigantional adjuvanted polyprotein HIV-1 vaccine in healthy
adults and effect of administered chloroquine. Clin. Vacc. Immunol. 21, 303–311.
Leroux-Roels, I., Forgus, S., De Boever, F., Clement, F., Demoitié, M.A., Mettens, P., Moris,
P., Ledent, E., Leroux-Roels, G., Ofori-Anyiman, O., The M72 study group, 2013.
Improved CD4+ T cell responses to Mycobacterium tuberculosis in PPD negative adults
by M72/AS01 as compared to the M72/AS02 and Mtb72F/AS02 tuberculosis can-
didate vaccine formulations: a randomized trial. Vaccine 31, 2196–2206.
Leroux-Roels, I., Koutsoukos, M., Clement, F., Steyaert, S., Janssens, M., Bourguignon, P.,
Cohen, K., Altfeld, M., Vandepapelière, P., Pedneault, L., et al., 2010. Strong and
persistent CD4+ T-cell response in healthy adults immunized with a candidate HIV-1
vaccine containing gp 120, Nef and Tat antigens formulated in three adjuvant sys-
tems. Vacine 28, 7016–7024.
Leroux-Roels, I., Leroux-Roels, G., Clement, F., Vandepapelière, P., Vassilev, V., Ledent,
E., Heineman, T.C., 2012. A phase 1/2 clinical trial evaluating safety and im-
munogenicity of a varicella zoster glycoprotein E subunit vaccine candidate in young
and older adults. J. Inf. Dis. 206, 1280–1290.
Levast, B., Awate, S., Babiuk, L., Mutwiri, G., Gerdts, V., van Drunen Littel-van den Hurk,
S., 2014. Vaccine potentiation by combination adjuvants. Vaccines 2, 297–322.
Lorent, J.H., Quetin-Leclercq, J., Mingeot-Leclercq, M.P., 2014. Amphiphilic nature of
saponins and their effects on artificial and biological membranes and potential
consequences for red blood and cancer cells. Org. Biomol. Chem. 12, 8803–8822.
Marciani, D.J., 2003. Vaccine adjuvants: role and mechanisms of action in vaccine im-
munogenicity. Drug Discov. Today 8, 934–943.
Marciani, D.J., 2018. Elucidating the mechanisms of action of saponin-derived adjuvants.
Trends Pharm. Sci. 39, 573–585.
Marty-Roix, R., Vladimer, G.I., Pouliot, K., Weng, D., Buglione-Corbett, R., West, K.,
MacMicking, J.D., Cheee, J.D., Wang, S., Lu, S., Lien, E., 2016. Identification of QS-21
as an inflammasome-activating molecular component of saponin adjuvants. J. Biol.
Chem. 291, 1123–1136.
Moris, P., Jongert, E., van der Most, R.G., 2018. Characterization of T-cell immune re-
sponses in clinical trials of the candidate RTS,S malaria vaccine. Hum. Vaccines
Immunother. 14, 17–27.
Olotu, A., Lusingu, J., Leach, A., Lievens, M., Vekemans, J., Msham, S., Lang, T., Gould, J.,
Dubois, M.C., Jongert, E., et al., 2011. Efficacy of RTS,S/AS01E malaria vaccine and
exploratory analysis on anti-circumsporozoite antibody titres and protection in
children aged 5-17 months in Kenya and Tanzania: a randomized controlled trial.
Lancet Infect. Dis. 11, 102–109.
Pasquier, F., Sadowsky, C., Holstein, A., Le Prince Leterme, G., Peng, Y., Jackson, N., Fox,
N.C, Ketter, N., Liu, E., Ryan, J.M., 2016. Two phase 2 multiple ascending dose
studies of vanutide cridificar (ACC-001) and QS-21 adjuvant in mild-to-moderate
Alzheimer's disease. J. Alzheimer Dis. 51, 1131–1143.
Penn-Nicholson, A., Geldenhuys, H., Burny, W., ven der Most, R., Day, C.L., Jongert, E.,
Moris, P., Hatherill, M., Ofori-Anyinam, O., Hanekom, W., The vaccine study team,
2015. Safety and immunogenicity of vaccine M72/AS01E in adolescents in a TB en-
demic setting. Vaccine 33, 4025–4034.
Reed, S.G., Orr, M.T., Fox, C.B., 2013. Key roles of adjuvants in modern vaccines. Nat.
Med. 19, 1597–1608.
RTS, S Clinical Trials Partnership, 2014. Efficacy and safety of the RTS,S/AS01 malaria
vaccine during 18 months after vaccination: a phase 3 randomized, controlled trial in
children and young infants at 11 African sites. PLoS Med. 11 (7), e1001685.
RTS, S Clinical Trials Partnership, 2015. Efficacy and safety of RTS,S/AS01 malaria
vaccine with or without a booster dose in infants and children in Africa: final results
of a phase 3, individually randomised, controlled trial. Lancet 386, 31–45.
Siegrist, C.A., 2013. Vaccine immunology. In: Plotkin, S.A., Orenstein, W.A., Offit, P.A.
(Eds.), “Vaccines”, Section 1, General aspects of Vaccination, 6th edition. Elsevier,
pp. 14–32.
Stassijns, J., Bollaerts, K., Baay, M., Verstraeten, T., 2016. A systematic review and meta-
analysis on the safety of newly adjuvanted vaccines amound children. Vaccine 7,
714–722.
Schwarz, T.F., Volpe, S., Catteau, G., Chlibek, R., David, M.P., Richardus, J.H., Lal, H.,
Oostvogels, L., Pauksens, K., Ravault, S., et al., 2018. Persistence of immune response
to an adjuvanted varicella-zoster virus subunit vaccine for up to year nine in older
adults. Hum. Vaccin. Immunother. 14, 1370–1377.
Van Braeckel, E., Bourguignon, P., Koutsoukos, M., Clement, F., Janssens, M., Carletti, I.,
Collard, A., Demoitié, M.A., Voss, G., Leroux-Roels, G., et al., 2011. An adjuvanted
polyprotein HIV-1 vaccine induces polyfunctional cross-reactive CD4+ T cell re-
sponses in seronegative volunteers. Clin. Infect. Dis. 52, 522–531.
Van den Berg, R., Coccia, M., Ripley Ballou, W., Kester, K.E., Ockenhouse, C.F.,
Vekemans, J., Jongert, E., Didierlaurent, A.M., van der Most, R.G., 2017. Predicting
RTS,S vaccine-mediated protection from transcriptomes in a malaria-challenge clin-
ical trial. Front. Immunol. 8, 557.
Van der Meeren, O., Hatherill, M., Nduba, V., Wilkinson, R.J., Muyoyeta, M., Van Brakel,
M.-A. Lacaille-Dubois
E., Ayles, H.M., Henostroza, G., Thienemann, F., Scriba, T.J., et al., 2018. Phase 2b
controlled trial of M72/AS01E vaccine to prevent tuberculosis. N. Engl. J. Med. 379,
1621–1634.
Vandepapelière, P., Horsmans, Y., Moris, P., Van Mechelen, M., Janssens, M., Koutsoukos,
M., Van Belle, P., Clement, F., Hanon, E., Wettendorff, M., Garçon, N., Leroux-Roels,
G., 2008. Vaccine adjuvant systems containing monophosphoryl lipid A and QS-21
induce strong and persistent humoral and T cell responses against hepatitis B surface
antigen in healthy adult volonteers. Vaccine 26, 1375–1386.
Vansteenkiste, J.F., Chul Cho, B., Vanakesa, T., De Pas, T., Zielinski, M., Kim, M.S.,
Jassem, J., Yoshimura, M., Dahabreh, J., Nakayama, H., et al., 2016. Efficacy of the
MAGE-A3 immunotherapeutic as adjuvant therapy in patients with resected MAGE-
A3-positive non-small-cell lung cancer (MAGRIT): a randomized, double-blind, pla-
cebo-controlled, phase 3 trial. Lancet Oncol. 17, 822–835.
Walkowicz, W.E., Fernández-Tejada, A., George, C., Corzana, F., Jiménez-Barbero, J.,
Ragupathi, G., Tan, D.S., Gin, D.Y., 2016. Quillaja saponin variants with central
glycosidic linkage modifications exhibit distinct conformations and adjuvant activ-
ities. Chem. Sci. 7, 2371–2380.
Welsby, I., Detienne, S., N'Kuli, F., Thomas, S., Wouters, S., Bechtold, V., De Wit, D.,
Gineste, R., Reinheckel, T., Elouahabi, A., Courtoy, P.F., Didierlaurent, A.M., Goriely,
S., 2017. Lysosome-dependent activation of human dendritic cells by the vaccine
12
Phytomedicine 60 (2019) 152905
adjuvant QS-21. Front. Immunol. 7, 663.
White, M.T., Bejon, P., Olotu, A, Griffin, J.T., Riley, E.M., Kester, K.E., Ockenhouse, C.F.,
Ghani, A.C., 2013. The relationship between RTS,S vaccine-induced antibodies,
CD4+ T cell responses and protection against Plasmodium falciparum infection. PLoS
One 8 (4), e61395.
Winblad, B., Graf, A., Riviere, M.E., Andreasen, N., Ryan, J.M., 2014. Active im-
munotherapy options for Alzheimer's disease. Alzheimer's Res. Ther. 6, 7.
World Health Organisation, 2016. Malaria vaccine: WHO position paper-January 2016.
Weekly Epidemiol. Rec. 91, 33–52.
www.ema.europa.eu/en/documents/overview/Shingrix-epar-medicine-overview_en.pdf.
Shingrix, INN-herpes zoster vaccine (recombinant, adjuvanted). EMA/76073/2018.
www.ema.europa.eu/en/documents/press-release/first-malaria-vaccine-receives-
positive-scientific-opinion-ema_en.pdf. EMA press release 2015. First malaria vaccine
receives positive scientific opinion from EMA, 2015. EMA/CHMP488348/2015.
www.who.int/campaigns/malaria-day/2018/en/. World Malaria Day, 25 April 2018.
www.who.int/malaria/media/malaria-vaccine-implementation-qa/en/. WHO/Q & A on
the malaria vaccine implementation programme (MVIP) January 2019.
www.who.int/news-room/fact-sheets/detail/hiv-aids. HIV/AIDS, Key facts, 19 July
2018.