MINIREVIEW

Vaccines against malaria—still a long way to go

Kai Matuschewski
Department of Molecular Parasitology, Institute of Biology, Humboldt University Berlin, Germany

Keywords Several species of Plasmodium cause a broad spectrum of human disease

gametocyte; immunization; malaria;
merozoite; Plasmodium falciparum;
Plasmodium vivax; sporozoite; vaccine
Correspondence
K. Matuschewski, Department of Molecular
Parasitology, Institute of Biology/Faculty of
Life Sciences, Humboldt University Berlin,
Philippstr. 13 #14, 10115 Berlin, Germany
Fax: +49 30 2093 6051
Tel: +49 30 2093 6053
E-mail: kai.matuschewski@hu-berlin.de
(Received 31 January 2017, revised 9 April
2017, accepted 10 May 2017)
doi:10.1111/febs.14107
that range from nausea and fever to severe anemia, cerebral malaria, and
multiorgan failure. In malaria-endemic countries, continuous exposure to
Plasmodium sporozoite inoculations and subsequent blood infections elicit
only partial and short-lived immunity, which gradually develops over many
years of parasite exposure and multiple clinical episodes. The ambitious
goal of malaria vaccinology over the past 70 years has been to develop an
immunization strategy that mounts protection superior to naturally
acquired immunity. Herein, three principal concepts in evidence-based
malaria vaccine development are compared. Feasible leads are typically
stand-alone subunit vaccine approaches that block Plasmodium parasite life
cycle progression or parasite/host interactions, and they constitute the
majority of candidates in preclinical research and early clinical testing.
Integrated approaches incorporate malaria antigen(s) into licensed or
emerging pediatric vaccine formulations. This strategy can complement the
malaria control portfolio even if the antimalarial component is only par-
tially effective and has led to the development of the only candidate vac-
cine to date, namely RTS,S-AS01. Experimental whole parasite vaccine
approaches have been repeatedly shown to elicit sterile and lasting protec-
tion against identical parasite strains, but mass production, proof of broad
protection against different parasite strains, and routes of vaccine delivery
remain significant translational road blocks. Global access to an effective
and affordable malaria vaccine will critically depend on innovative transla-
tional research that builds on a better molecular understanding of Plasmod-
ium biology and host immunity.

Introduction
Malaria remains the most important mosquito-borne
infectious disease and is caused by blood infection
with Plasmodium parasites [1]. These obligate intracel-
lular pathogens belong to the phylum Apicomplexa,
which are single-cell eukaryotes that coevolved with
their respective hosts across the entire animal kingdom
[2]. What distinguishes Plasmodium is their unidirec-
tional population expansion first in the liver and then
in red blood cells before differentiation into sexual
stages that colonize Anopheles mosquitoes [3].
Plasmodium stage conversion over the course of the
human infection offers multiple targets for vaccine-
induced immunity (Fig. 1), but the parasite is tailored to

Abbreviations
BCG, Bacillus Calmette-Gue
rin; CelTOS, cell-traversal protein for ookinetes and sporozoites; ChAD63, chimpanzee adenovirus 63; CSP,
circumsporozoite protein; GLURP, glutamate rich protein; GMO, genetically modified organism; MSP, merozoite surface protein; MVA,
modified vaccinia Ankara virus; PfEMP1, Plasmodium falciparum erythrocyte membrane protein 1; RTS,S/AS01, proprietary vaccine
formulation by GlaxSmithKline Biologicals; SSP, sporozoite surface protein; TRAP, thrombospondin-related anonymous protein.

The FEBS Journal (2017) ª 2017 Federation of European Biochemical Societies 1

Malaria vaccine perspectives K. Matuschewski

reinfect the host lifelong. Naturally acquired immunity
develops only gradually after > 20 clinical episodes over
the course of many years [4], but it is never sterile [5]. Nat-
ural immunity mostly comprises plasma B cells that pro-
duce antibodies against variant parasite-encoded
erythrocyte surface antigens, such as Plasmodium falci-
parum erythrocyte membrane proteins (PfEMP1s) and, to
a lesser extent, merozoite proteins (Fig. 1) [5]. In marked
contrast, acute infections by viral and some bacterial
pathogens elicit lasting and frequently lifelong protection
against reinfection, often after a single infection. This
immunological memory is the basis for protective vaccines
[6] and clearly missing in malaria and other chronic infec-
tious diseases, such as tuberculosis. Accordingly, simply
extrapolating successful antimicrobial vaccines to malaria
is likely inadequate, and despite intense efforts in malaria
vaccinology over the past 70 years the goal to develop an
effective malaria vaccine that performs superior to natu-
rally acquired immunity has not yet been achieved.
A hypothetical malaria vaccine is highly unlikely to
replace existing malaria control tools, unless it achieves
complete sterilizing immunity. Rather a vaccine will be
an integral part of malaria control programs, which
include access to rapid diagnosis and treatment with
Artemisinin-based combination therapies, ownership
of long-lasting insecticide treated nets, and regular
campaigns of indoor residual spraying with a suitable
insecticide. Malaria control has been successful over
the past 10 years because of exceptional donor com-
mitment. Continuation of effective malaria manage-
ment and further reduction in the malaria disease
burden in high-risk endemic areas critically depends
on sustained recognition of malaria as a global health
priority. Furthermore, widespread tolerance against
insecticides in Anopheles vector populations and para-
site resistance against every approved antimalarial
drug on the market emphasize the urgent need for a
safe and effective vaccine.

Fig. 1. Adaptive immune responses during Plasmodium life cycle progression in humans. (1) During an infectious Anopheles bite
sporozoites (green) are injected and migrate through the skin to enter a blood capillary and eventually invade a suitable hepatocyte.
Plasma cells (purple) originating from activated memory B cells and secreting antibodies against SSPs can immobilize sporozoites and
reduce the infectious parasite dose. (2) During the first pre-erythrocytic asexual replication phase in the liver a single successful
sporozoite forms thousands of infectious merozoites. Cytolytic effector memory CD8+ T cells (T) can specifically recognize and lyse
infected hepatocytes. Merozoites initiate the second asexual replication phase inside erythrocytes. (3) Plasmodium falciparum, unlike
other Plasmodium species, extensively remodels the erythrocyte surface resulting in sequestration of infected erythrocytes to the
vascular endothelium. This so-called cytoadherence is a major cause of severe disease. A large range of plasma cells (yellow) originating
from activated memory B cells and secreting antibodies against the P. falciparum-encoded surface protein repertoire correlates with
gradual acquisition of naturally acquired immunity in malaria-endemic areas. (4) Plasma cells (blue) originating from activated memory B
cells and secreting antibodies against MSPs can immobilize merozoites and reduce parasitemia in the blood circulation. A subset of
merozoites differentiates into female (blue) and male (pink) gametocytes, which may be ingested by an Anopheline mosquito and lead to
completion of the life cycle.

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K. Matuschewski
Vaccine opportunities to break the
Plasmodium cycle
Parasitic eukaryotes are complex pathogens and some
of the finest artists in escaping host defense mecha-
nisms, including adaptive immune responses [7,8]. No
correlates of protection against malaria, that is,
immune signatures or antigens that permit distinction
between previous parasite exposures vs. protective
immunity, have been identified yet [5,9]. Therefore,
malaria vaccine development still relies on empirical
testing in preclinical models [10] and small human
challenge studies [11]. Four principal, and ideally com-
plementary, biological targets, namely sporozoites,
liver stages, and asexual and sexual blood stages
(Fig. 1), can be exploited for vaccine development to
fast-forward naturally acquired immunity to curtail—
in the order of priority—(a) severe disease, (b) clinical
malaria episodes, (c) parasite load, and (d) community
transmission [12–16].
Sporozoites are the infectious agents that are
injected during the probing phase of an infectious
Anopheles bite (Fig. 1). They actively migrate through
the skin in search of a blood capillary to be flushed to
the final target organ, the liver [3,17]. Antibody-
mediated inhibition of sporozoite infection of the liver
can reduce the initial parasite inoculum and requires
the induction of long-lived antibody responses [18].
Accordingly, antisporozoite vaccines should stimulate
the formation of germinal center B cells, which can
maintain an effector phenotype and initiate rapid
plasma cell differentiation through interaction with
memory T follicular helper cells [19]. However, also in
the secondary response, upon parasite antigen rechal-
lenge, high levels of inhibitory antibodies are only
formed within a few days. In the meantime, the sporo-
zoite is already in full liver stage development. Accord-
ingly, antisporozoite vaccine strategies that elicit
antibodies can only achieve substantial reduction in
Plasmodium transmission in subsequent infections and
have an impact over time. However, they consistently
fail in preventing malaria infections [20] and, hence,
are not considered in travel medicine.
During the obligate pre-erythrocytic phase in the
liver, Plasmodium undergoes an enormous population
expansion resulting in the formation of thousands of
first-generation merozoites (Fig. 1). Only during liver
stage maturation is the parasite susceptible to direct T-
cell-mediated killing, and this period is long enough
(~ 7 days) for secondary recall responses [3,21]. Last-
ing and robust antiliver stage immunity is the principal
mechanism for the experimental success of vaccine
strategies that lead to live, but developmentally
The FEBS Journal (2017) ª 2017 Federation of European Biochemical Societies
Malaria vaccine perspectives
arrested, liver stages [22–25]. Although work in murine
malaria models has consistently recovered antigen-spe-
cific cytolytic effector memory CD8+ T cells as the
main cell type that recognizes and lyses infected hepa-
tocytes [26–29], a similar important role remains to be
shown for human malaria.
Merozoites are tailor-made to actively and rapidly
invade red blood cells (Fig. 1). This second asexual
replication phase inside erythrocytes is the exclusive
cause of malaria and an ideal target to reduce the inci-
dence of clinical attacks. However, antibody-mediated
inhibition and destruction of merozoites [30] is limited
to a very brief (~ 30 s) moment, when the parasite is
outside its host cell and in the process of invading a
new erythrocyte. Many candidate antigens that per-
form vital functions in the invasion process are stored
in specialized organelles, termed micronemes, rhop-
tries, and dense granules, and are sequentially released
upon intimate merozoite contact with the erythrocyte
surface, thus further limiting exposure to inhibitory
antibodies [31]. A population-based study of antibody
responses in malaria-endemic countries did not return
any signatures of protection, and many candidate anti-
gens were recognized to a similar extent by protected
and susceptible individuals [32]. In addition, increasing
evidence suggests that memory B cells, which are the
principal antibody-secreting cells that target blood
stage antigens upon vaccine-induced immunity, appear
to contract remarkably early [33].
Severe malaria in a proportion of children infected
with P. falciparum remains a leading cause of death in
Afrotropical regions [34]. After invasion, P. falciparum
extensively refurbishes the host erythrocyte and
exports PfEMP1s and other variant antigens to the
surface. Binding of infected erythrocytes to cognate
receptors on the vascular endothelium, a process ter-
med cytoadhesion, is crucial to avoid splenic clearance
and results in a large pathological spectrum, ranging
from capillary occlusion and hypoxia to tissue inflam-
mation and multiorgan failure [35]. A broad range of
memory B cells that target the P. falciparum-encoded
surface protein repertoire is gradually acquired during
infancy and childhood and reflects the slow acquisition
of antidisease immunity in malaria-endemic areas.
Whether other human Plasmodium species, including
Plasmodium vivax, express parasite-encoded surface
markers that can be targeted by inhibitory antibodies
remains an unsolved matter.
A proportion of merozoites eventually differentiate
into sexual stages, which can recombine and complete
the life cycle upon ingestion by an Anopheles mos-
quito. Although of no immediate benefit to the vacci-
nated individual high-titer antibodies against
3
Malaria vaccine perspectives
gametocytes and the diploid zygote might aid in reduc-
tion of Plasmodium transmission on a community-level
[36]. However, when prioritizing a small subset of can-
didate antigens for the most vulnerable population,
infants and pregnant women, gamete and ookinete sur-
face proteins are negligible.
Together, two of the five parasite stages in the mam-
malian host, namely liver stages and parasite antigens
on the infected erythrocyte surface, represent promis-
ing targets for vaccine strategies. Liver stages can be
targeted in all Plasmodium species and an effective vac-
cine can elicit complete protection against blood infec-
tion. But how to accurately induce a range of effector
memory CD8+ T cells that home to the liver upon re-
infection remains a principal problem in vaccinology
[37]. Antibody-mediated inhibition of P. falciparum-
encoded erythrocyte surface proteins can prevent dis-
ease and targets the long replication phase inside red
blood cells. But management of antigenic variation is
a persistent challenge. On the other hand, induction of
antibody-mediated inhibition of sporozoite and mero-
zoite invasion or gamete fusion in the mosquito mid-
gut is less demanding. While either one may offer
synergistic benefits to an antiliver stage or anticytoad-
hesion vaccine, none are likely to curtail Plasmodium
life cycle progression on their own. It follows that
malaria vaccine research aims at developing a multi-
component, multistage vaccine, similar to pediatric
combination vaccines against multiple childhood
diseases.
Feasible leads: testing one antigen at
a time
Until date, the vast majority of candidate antigens are
designed as stand-alone malaria subunit vaccines
(Table 1). Major advantages of a distinct antigen for-
mulation include the potential feasibility to produce a
defined malaria vaccine under good manufacturing
procedures, at reasonable cost, and in a formulation
that is thermostable and easy to deliver intramuscu-
larly to human subjects. Accordingly, identification of
a protective antigen and its translation to a safe and
protective malaria subunit vaccine would be the most
desirable solution. Over the past 30 years, a large
range of candidate antigen formulations often included
parasite antigens in the historic order of recombinant
cloning. They encompassed initially the major sporo-
zoite surface proteins (SSPs) circumsporozoite protein
(CSP/SSP1) and thrombospondin-related anonymous
protein (TRAP/SSP2), the major merozoite surface
proteins 1–3 (MSP1–3), and the sexual stage antigens
25 and 230 (s25, s230). However, experimental proof
4 The FEBS Journal (2017) ª 2017 Federation of European Biochemical Societies
K. Matuschewski
that any of the Plasmodium antigens is a signature of
protective immunity as opposed to parasite exposure
remains elusive [32].
A landmark study established that a recombinant
malaria vaccine, comprising MSP1, MSP2, and ring-
infected erythrocyte surface antigen, could exert selec-
tive pressure on parasite populations [38]. Although
this finding signifies selective immunity against some
antigens, in this case MSP2, it highlights swift parasite
evasion of vaccine-induced selection pressure. A recent
completion of the largest field trial of a blood stage
vaccine candidate formulation, consisting of recombi-
nant MSP3 and glutamate-rich protein (GLURP) in
alum, in 1,735 children revealed no efficacy [39], reiter-
ating enormous challenges for merozoite vaccine
approaches. These examples illustrate that antigenic
variation and polymorphic antigens inevitably remain
the largest obstacles for subunit vaccine development.
Only informed design of a multivalent vaccine incor-
porating several invariant antigens can surmount this
persisting challenge.
An alternative approach is the delivery of recombi-
nant virus particles to express Plasmodium antigens, in
this case TRAP/SSP2 [40]. Multiple rounds of opti-
mization returned a heterologous prime/boost protocol
with chimpanzee adenovirus 63 (ChAd63) and modi-
fied vaccinia Ankara virus (MVA), respectively. A
recent small phase II study with 115 participants in
Senegal showed no efficacy of this vaccine, and com-
bined analysis with earlier data from Kenia returned
only a small, albeit significant, delay until patency [40].
A number of candidate antigens are still in the
exploratory, preclinical phase or phase I safety testing.
One example is a formulation with the cell-traversal
protein for ookinetes and sporozoites (CelTOS), which
was tested in an elegant murine infection model using
transgenic chimeric parasites that express the P. falci-
parum protein [41]. Efficacy was very modest leading
only to a twofold reduction in parasite liver load,
which does not translate in any detectable effect on
Table 1. Exemplary candidate subunit preparations.
Target antigen Delivery platform Reference
Phase II clinical trials
GLURP, MSP3 Protein/alum [39]
TRAP ChAd63/MVA [40]
Phase I trials/ preclinial
CelTOS Protein/AS10B adjuvant [41]
Var2-CSA Protein/alhydrogel [42]
s25, s230 ChAd63/MVA [43]
s25, s230, sexual stage antigen 25, 230.
K. Matuschewski
blood infection. Therefore, such a vaccine needs to be
considerably re-engineered before any further testing,
including in clinical trials. An attractive candidate is
the VAR2CSA surface protein on infected erythro-
cytes, which mediates binding to chondroitin sulfate
A (CSA) on the placenta and is a major factor for
placental malaria in pregnant women [42]. Many
obstacles remain, but the development of a tailored
vaccine for young women will be an important step-
ping-stone toward anticytoadhesion vaccines. Finally,
sexual stage antigens to induce transmission-blocking
immunity can be tested in membrane feeding assays
that partly mimic natural transmission to Anopheline
mosquitoes. A systematic analysis returned two anti-
gens, s25 and s230, as lead candidates for further
investigation [43]. Together, feasible subunit vaccine
approaches often deliver disappointing results in effi-
cacy testing in the field, and conserved, nonredun-
dant antigens that are indispensable for parasite life
cycle progression need to be targeted to avoid
counterselection [38].
Integrated vaccines: taking advantage
of existing pediatric vaccines
As a valuable addition to sustain affordable malaria
control, a candidate vaccine needs to perform safely,
and ideally efficiently, during neonatal and early life
vaccination in resource-poor settings. Accordingly,
such a malaria childhood vaccine needs to be tailored
for integration into the expanded program of immu-
nization. A powerful answer is to swing aboard
licensed pediatric vaccines and add-on a malaria anti-
gen. When combined as prime-boost strategy employ-
ing various vaccine approaches Plasmodium infections
may be targeted in addition to the original pathogen
(Table 2). Most importantly, because they are not
stand-alone formulations integrated malaria vaccines
are not required to perform to perfection from the
beginning and can be improved iteratively, while
always maintaining their original purpose, for instance
complete protection against hepatitis B.
It is, therefore, no surprise that only one candidate
malaria vaccine, which followed this approach from
the original design, ever advanced to pivotal phase III
clinical trials, namely RTS,S/AS01 [44]. This vaccine
formulation resembles an advanced hepatitis B vac-
cine, which was the world’s first recombinant vaccine
[45]. The vaccine consists of virus-like particles that
can be produced in large fermenters by recombinant
expression of the hepatitis B surface (S) antigen in
Saccharomyces cerevisiae. The modified version still
contains the S antigen as the largest protein portion,
The FEBS Journal (2017) ª 2017 Federation of European Biochemical Societies
Malaria vaccine perspectives
Table 2. Integrated vaccination approaches.
Integrated Plasmodium
Original vaccine version antigen Reference
Phase III clinical trials
Hepatitis B RTS,S/AS01 PfCSP fragment [44,47,48]
vaccine
Preclinical evaluation
Yellow fever YF17D/PyCS PyCSP fragment [49]
17D
BCG Glaxo BCG-CS PfCSP full length [50]
Pf, Plasmodium falciparum; Py, Plasmodium yoelii.
but in addition a smaller fraction consisting of a por-
tion of the P. falciparum CSP, including the repeat
region (R) and a T-cell epitope (T), fused to the S
antigen. After multiple rounds of safety, immuno-
genicity and efficacy testing and improvements of a
formulation with a potent liposome-based adjuvant,
termed AS01 [46], was selected for large-scale studies
in at-risk populations [47]. In all studies, there was an
initial short phase of protection against clinical malar-
ia, in good correlation with exceptionally high anti-
body titers resulting from the vaccination scheme. As
expected [20], protection disappeared with the concur-
rent decrease in antibody titers after few months.
Since initial protection was offset in later years the
overall result was no efficacy [48]. These disappointing
results reflect the fundamental differences in infection
biology between viral and parasitic pathogens and
cast doubt whether integrated approaches work in
principle.
Nevertheless, the importance of the RTS,S/AS01 tri-
als cannot be overestimated. They achieved the first
advancement of a candidate vaccine against any of the
three most important infectious diseases, HIV/AIDS,
tuberculosis, and malaria, to phase III clinical trials,
the set-up of clinical trials capacity in malaria-ende-
mic countries, and a robust foundation for further
developments. Important research topics are to
uncover whether (a) CD4+ and CD8+ T-cell
responses were induced in addition to antibody
responses, (b) the selected Plasmodium antigen is cen-
tral to pre-erythrocytic immunity, (c) alternative liver
stage antigens might perform superior, and (d)
biomarkers other than antibody titers might be better
predictors of vaccine efficacy.
It will also be interesting to explore whether combi-
nation with other integrated formulations can improve
the outcome (Table 2). A recombinant yellow fever
vaccine has been designed for preclinical studies in the
murine malaria model Plasmodium yoelii [49], a per-
haps critical intermediate step toward translation,
5
Malaria vaccine perspectives K. Matuschewski

which was omitted entirely in the development of Early proof of principle studies demonstrated sterile
RTS,S/AS01 [10]. The yellow fever vaccine is based on immunity in humans that were immunized by exposure
a live, attenuated virus and a single dose protects for to c-irradiated P. falciparum-infected mosquitoes, and
life. It will be interesting to explore potential protec- these findings could be consistently repeated [51]. Mos-
tion against malaria once a suitable P. falciparum anti- quito delivery of c-irradiated sporozoites (cspz) was
gen is selected. The recombinant expression of soon recognized unrealistic, but these studies were an
P. falciparum CSP in the childhood tuberculosis vac- impressive testimonial for the validity of the original
cine strain Bacillus Calmette-Gu
erin (BCG) is an inter- concept [22], the important contribution of murine
esting research direction, since this approach builds on malaria models for evidence-based malaria vaccine
the robust safety and efficacy of the only live, attenu- development [10], and, perhaps most critical, the feasi-
ated antibacterial pediatric vaccine [50]. Immunity bility to develop a malaria vaccine on the basis of
against the complex intracellular bacterium Mycobac- arrested liver stage development.
terium tuberculosis is likely to resemble anti-Plasmodium Limitations of the cspz production include strict
immunity much closer than antiviral immunity and also dependence on biosafety level 3 facilities and batch-to-
requires only one immunization dose. However, experi- batch variations, because c irradiation induces uncon-
mental testing of vaccine safety and efficacy proves diffi- trolled DNA double-strand breaks. Moreover, the full
cult, since a preclinical animal model was not range of liver stage-induced immunity may be missed,
considered [50]. Other pediatric vaccines that are because cspz arrest very early in liver infection, imme-
routinely delivered, including against measles, might diately after sporozoite transformation [21]. A strong
also serve as suitable platforms for integrated malaria advantage of the cspz technology is the universal cov-
vaccine strategies. erage of every human Plasmodium species and the abil-
ity to implement multiple vaccine strains, which can
cover regional parasite diversity in the tropics [52].
Attenuated liver stages: benchmark
Some of the restrictions of cspz were overcome by
experimental malaria vaccines
targeted removal of stage-specific Plasmodium genes
In recognition of the complex molecular make-up and that perform vital functions for liver stage maturation
intriguing escape mechanisms of Plasmodium parasites [24]. Genetically arrested parasites (GAPs) can now be
as well as the persistent challenges to subunit malaria engineered as precision malaria vaccines, tailored to
vaccines, major efforts now return to the origins of arrest at any given point in pre-erythroyctic develop-
malaria vaccine development [22]. The unifying princi- ment and harboring additional components, such as
ple is to elicit lasting, sterile immunity by immuniza- blood stage antigens and ligands to stimulate antigen
tion with live Plasmodium sporozoites, which cannot presentation. Of note, occupational exposure or acci-
progress to blood infection (Table 3). dental mosquito release no longer pose health risks,
greatly facilitating potential production in malaria-
endemic countries. P. falciparum GAPs have already
Table 3. Live, metabolically active Plasmodium sporozoite advanced to clinical testing, and first safety and
inoculations. immunogenicity data are encouraging [53]. However,
in marked contrast to cspz, GAP technology requires
Approach Advantages Challenges Reference
re-engineering by reverse genetics for every Plasmod-
In clinical trials ium species and strain.
Chloroquine Superior immunity Patient [23,54] An alternative approach to stop blood infection is
cover compliance sporozoite delivery under simultaneous drug cover
c-Irradiation Applicable to all Low immunity [22,51]
[21,23,25]. Again, the success of a preclinical model,
(cspz) species and strains
where mice were given normal sporozoites together
Gene deletion Precision liver GMO strains [24,53]
(GAP) stage arrests, with the antimalarial drug chloroquine and enjoyed
no BSL3 facility lasting protection against challenge infection [23], were
required repeated successfully in small human trials [54]. Drug
Preclinical validation cover can also be done with the antibiotic azithromy-
Azithromycin Superior immunity Patient [25] cin, which leads to release of attenuated liver stage
cover compliance
merozoites and superior protection [25], and the 8-ami-
Primaquine Elimination of Low immunity [55]
noquinoline primaquine, which eliminates developing
cover liver stages
and quiescent liver stages [55] (Table 3). Major safety
BSL3, biosafety level 3; GMO, genetically modified organism. concerns with all drug cover approaches are patient

6 The FEBS Journal (2017) ª 2017 Federation of European Biochemical Societies

K. Matuschewski
compliance and a range of pharmacokinetics leading
to inconsistent liver stage arrest in different individu-
als.
Whole parasite vaccines: terminal or
roadmap from bench to field?
Four significant hurdles need to be overcome before
whole parasite vaccines can be reasonably implemented
in malaria-endemic countries. (a) Mass rearing of
infected Anopheles mosquitoes, hand-dissection of sali-
vary glands, and sporozoite purification are labor-
intense processes. Broad access and affordability will
critically depend on replacement by automated mos-
quito-free, axenic sporozoite culture systems. (b) Wide-
ranging protection against different parasite strains is
vital and was addressed only very recently. Protection
against challenge with a heterologous P. falciparum
strain was incomplete in subjects immunized by three
doses of sporozoite inoculations under chloroquine
cover [56] and negligible in individuals immunized with
five high doses of Pfcspz [57]. These unsatisfactory
results raise the alarming, yet plausible, possibility that
robust sterile protection elicited by whole parasite vacci-
nes is not strain transcending. If confirmed, a potential
solution could be vaccine cocktails containing different
strains, but selection for vaccine-resistant strains could
emerge as a major risk. (c) Vaccines need to be delivered
safely by trained field workers, and the preferred route
of vaccine delivery is intramuscular or subcutaneous
needle injection. Thus far, lasting protection was only
achieved by intravenous sporozoite injection, which is
an unacceptable administration route for preventive
vaccines. (d) As with all eukaryotic cells, long- and
short-term storage of viable parasites needs to be done
in liquid nitrogen and dry ice, respectively. Since main-
taining a cold chain for vaccine delivery in the tropics is
already an enormous logistic challenge, deep-frozen
chains to some of the world’s most remote communities
seem too visionary, and alternative long-term ther-
mostabilization techniques need to be developed.
To overcome all four roadblocks and reroute the
development of whole sporozoite vaccines from a
deadlock to a convincing road map, extensive bioengi-
neering and basic science investments have to be made.
But the complex infection biology of Plasmodium para-
sites might require complex solutions, such as whole
parasite vaccines.
Conclusion and outlook
In the past malaria vaccinology has focused on con-
cepts that were developed against acute infections with
The FEBS Journal (2017) ª 2017 Federation of European Biochemical Societies
Malaria vaccine perspectives
viral and bacterial pathogens. Eliciting strong recall
antibody responses alone appear insufficient against
Plasmodium parasites. These eukaryotic cells evolved
sophisticated strategies to escape invasion inhibition
by sequential secretion of redundant parasite ligands
and to overcome recognition of visible antigens by
antigenic diversity. Integration of candidate Plasmod-
ium antigens into licensed vaccines for neonatal and
childhood immunization remains largely underex-
plored. The historic momentum gained by multicenter
clinical trials of RTS,S/AS01 should be utilized to
advance this vaccine further and initiate immunologi-
cal studies in murine malaria models to gain mechanistic
insights into the potential effects against invading sporo-
zoites and developing liver stages, which remain inacces-
sible in human subjects. Since the choice of target antigen
is essential a better molecular understanding of what dis-
tinguishes a protective immune response from immune
recognition of parasite exposure is important. For any
subunit vaccine approach the necessary evidence to
advance to clinical testing should be based on robust
indications for signatures of protection. Whole sporo-
zoite vaccines need to be 100% effective to be successful,
but can also provide the missing roadmap and assist in
prioritizing Plasmodium pre-erythrocytic antigens.
Pioneering work beyond classical vaccine development,
readiness of each stakeholder to start all over again, and
public awareness of one of the most important challenges
in medicine are vital to further accelerate malaria vacci-
nology. Yet, the discovery, development, and licensing of
a malaria vaccine formulation, which meets all necessary
requirements, including safety, affordability, accessibility,
applicability, and efficacy, is far beyond the horizon.
Acknowledgements
Diane Schad (Max Planck Institute for Infection Biol-
ogy) designed the illustration. Apologies to the many
researchers in this field whose work could not be cited
directly in this mini-review due to space constrains.
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