MINIREVIEW
Multifunctional hostdefensepeptides: antiparasitic
activities
Amram Mor
Department of Biotechnology & Food Engineering, Technion – Israel Institute of Technology, Haifa, Israel
Introduction
Human parasites are responsible for millions of deaths
around the world every year. Malaria, for instance, is
endemic in over 100 Third World countries, with an
estimated 400 million clinical cases correlating with
1–3 million deaths annually, and over 3 billion inhabit-
ants of tropical regions are considered to be at risk [1].
Of particular concern is the causative agent of human
malaria, Plasmodium falciparum, a large number of
strains of which are drug resistant, in particular to
chloroquine [2]. Reports of field strains of P. falcipa-
rum demonstrating in vitro resistance to artemisinins –
recently introduced antimalarial drugs – are also
alarming [3]. Similarly, leishmaniases are important
causes of morbidity and mortality in humans and ani-
mals on four continents, and are extremely difficult to
treat [4]. There is thus a clear need for new therapeutic
agents against these and related parasites. A multitude
of preliminary studies suggest that hostdefense pep-
tides (HDPs) represent a promising route towards
developing new, efficient antiparasitic therapies [5].
Magainins and cecropins were among the very first
examples of animal HDPs reported to display antipar-
asitic activities, some 20 years ago [6], and synthetic
hybrids of cecropin and melittin have exhibited
enhanced potency [7]. These notorious antimicrobial
peptides are known to function as part of an inducible
immune response against a number of microbial infec-
tions. Thus, when injected into anopheline mosquitoes
previously infected with a variety of Plasmodium spe-
cies, the antimicrobial peptides disrupted sporogonic
Keywords
antimicrobial peptide; chemical mimicry;
drug design; drug resistance; infectious
disease; Leishmaniasis; malaria; membrane
active compound; oligo-acyl-lysyl;
peptide-mimetic
Correspondence
A. Mor, Laboratory of Antimicrobial Peptides
Investigation (LAPI), Department of
Biotechnology & Food Engineering,
Technion – Israel Institute of Technology,
Haifa, Israel
Fax: +972 4 829 3399
Tel: +972 4 829 3340
E-mail: amor@tx.technion.ac.il
(Received 28 April 2009, revised 12 August
2009, accepted 12 August 2009)
doi:10.1111/j.1742-4658.2009.07358.x
Whereas significant knowledge is accumulating on the antibacterial and
antifungal properties of hostdefense peptides (HDPs) and their synthetic
mimics, much less is known of their activities against parasites. A variety
of in vitro and in vivo antiparasitic assays suggest that these notorious anti-
microbial compounds could represent a powerful tool for the development
of novel drugs to fight parasites in the vertebrate host or to complement
current therapeutic strategies, albeit the fact that HDPs essentially act by
nonspecific mechanisms casts serious doubt on their ability to exert suffi-
cient selectivity to be considered ideal candidates for drug development.
This minireview summarizes recent efforts to assess the antiparasitic prop-
erties of HDPs and their synthetic derivatives, focusing on two of the most
used models – Plasmodium and Leishmania species – for antiparasitic
assays against the different development stages.
Abbreviations
APP, antilipopolysaccharide factor; HDP, hostdefense peptide; Hst5, histatin-5; OAK, oligo-acyl-lysyl.
6474 FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS
development by aborting the normal development of
oocysts. As the vector cannot transmit the parasite to
another host, this suggested the possibility of induc-
ing effective transmission-blocking immunity in the
mosquito vector by transgeneic expression of genes
coding for magainins, cecropins or similarly acting
parasiticidal peptides in the mosquito genome [6].
Since then, antiparasiticactivities have been reported
for numerous additional antimicrobial peptides and
their synthetic derivatives. Representative antiparasitic
peptides (APPs) are listed in Table 1, and reports from
the past decade are briefly outlined below.
APPs
Among the structurally constrained peptides, defensins
are by far the most investigated family of HDPs, and
numerous members have been reported to exert a vari-
ety of antiparasitic activities. For instance, the classic
antifungal defensin, drosomycin, was shown to signifi-
cantly inhibit the development of Plasmodium berghei
ookinetes at micromolar concentrations [8]. Scorpine,
a 75-mer peptide from scorpion venom Pandinus impe-
rator whose structure resembles a hybrid between
defensin and cecropin, was reported to inhibit both the
Table 1. List of representative natural and synthetic antiparasitic peptides. MIC and IC
50
: minimal concentrations inducing 100% and 50%
inhibition of parasite growth, respectively. MLC and LC
50
: minimal concentrations inducing 100% and 50% lysis of the parasite, respectively.
Compound Primary structure Parasite
Antiparasitic
effect (l
M) Reference
Magainin II GIGKFLHSAKKFGKAFVGEIMNS Paramecium caudatum MIC = 4.1 36
Tetrahymena pyriformis MIC = 8.1
Acanthamoeba castellani MIC = 0.9
Cecropin–melittin KWKLFKKIEKVGQGIGAVLKVLTTGL P. falciparum IC
50
10 7
D-HALO-rev AKKLOHALHOALLALOHLAHOLLAKK P. falciparum IC
50
= 0.1 24
Hst5 DSHAKRHHGYKRKFHEKHHSHRGY Leishmania donovani LC
50
= 7.3 26
Leishmania pifanoi LC
50
= 14.4
NK-2 KILRGVCKKIMRTFLRRISKDILTGKK T. cruzi IC
50
= 2.5 15
P. falciparum LC
50
= 6.2 14
Angiotensin II NRVYIHPF P. gallinaceum IC
50
30 16
aI-Defensin ACYCRIPACIAGERRYGTCIYQGRLWAFCC T. cruzi IC
50
937
hII-Defensin Cyclic GVCRCLCRRGVCRCLCRR Leishmania amazonensis IC
50
12.5 11
Drosomycin DCLSGRYKGPCAVWDNETCRRVCKEEGR
SSGHCSPSLKCWCEGC
P. berghei IC
50
10 8
Gomesin ZCRRLCYKQRCVTYCRGR P. falciparum IC
50
80 22
P. berghei IC
50
50
Buforin II TRSSRAGLQFPVGRVHRLLRK C. parvum IC
50
20 20
Bombinin H4 I
IGPVLGLVGSALGGLLKKI L. donovani IC
50
= 1.7 17
L. pifanoi IC
50
= 5.6
Temporin A FLPLIGRVLSGIL L. donovani LC
50
= 8.4 18
L. pifanoi LD
50
12
Tachyplesin KWCFRVCYRGICYRRC L. braziliensis LD
50
= 4.7 21
T. cruzi LD
50
= 87.8
Tyrothricin A Cyclic VOLFPFFNQY P. falciparum IC
50
= 0.0006 27
Gramicidin S Cyclic VOLFPVOLFP P. falciparum IC
50
= 1.3 27
K
4
K
20
S4 ALWKTLLKKVLKAAAKAALKAVLVGANA P. falciparum IC
50
= 0.2 30
K
4
S4(1–13) ALWKTLLKKVLKA P. falciparum IC
50
=6 31
L. major MLC = 12.5 28
C
3
-K
4
S4(1–13) Propanoyl-ALWKTLLKKVLKA P. falciparum IC
50
=4 31
L. major MLC = 18 28
NC
7
-K
4
S4(1–13) Aminooctyl-ALWKTLLKKVLKA L. major MLC = 6.25 28
P. falciparum IC
50
532
Dermaseptin O1 GLWSTIKQKGKEAAIAAAKAAGQAALGAL T. cruzi MLC = 2.8 38
L. amazonensis MLC = 23 39
OAKs
a
lK-LK-LK-LK P. falciparum IC
50
= 0.08 35
LK-LK-LK P. falciparum IC
50
= 0.14
a
Acyl-lysyl oligomers where: l, dodecanoyl; L, aminododecanoyl; K, lysyl. Underlined letter designates amino acids in the D-configuration.
A. Mor Antiparasitic peptides
FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS 6475
ookinete and gamete developmental stages of Plasmo-
dium berghei with ED
50
values of 0.7 and 10 lm,
respectively [9]. There are newer reports of transgenic
scorpine producing 98% mortality in the sexual
stages of P. berghei at 15 lm and a 100% reduction in
P. falciparum parasitemia at 5 lm [10]. Various defen-
sins also have leishmanicidal properties, possibly by
inducing apoptosis [11]. The fact that substantially
higher leishmanicidal activity was observed against
mutant strains of Leishmania major in which leishman-
olysin, a surface metalloprotease, was knocked out
suggests that virulence factors such as leishmanolysin
might prevent antimicrobial peptide-mediated apop-
totic killing. Longicin, another member of the defensin
family, from the tick Haemaphysalis longicornis, exhib-
ited antiparasitic activity in vitro and in vivo against
the erythrocyte stage of Babesia, the merozoites, by
preventing or retarding their proliferation. The fact
that longicin exerted a hemoparasiticidal effect without
demonstrable toxicity against mammalian host cells
suggests its usefulness as a model for the development
of chemotherapeutic compounds against tick-borne
disease organisms [12]. A recent report studied clinical
isolates from three microsporidia species, using spore
germination and enterocyte-like cell infection assays
to determine the effect of a panel of antimicrobial
peptides: lactoferrin, lysozyme, human b-defensin-2,
human a-defensin-5, and human a-defensin-1. The
antimicrobial peptides have been reported to efficiently
inhibit spore germination and ⁄ or cell infection of one
or a number of isolates, either alone or upon combina-
tion with lysozyme, suggesting that intestinal anti-
microbial peptides contribute to the prevention of
infection by luminal microsporidia spores [13].
The linear 27-residue synthetic derivative of the
porcine protein NK-lysin, NK-2, was found to exert
selective activity against P. falciparum [14]. Infected
human erythrocytes were rapidly permeabilized by
NK2 at 5–10 lm, which reduced the viability of the
intracellular parasite, whereas noninfected cells were
hardly affected. This selectivity was ascribed to loss
of plasma membrane asymmetry and concomitant
exposure of phosphatidylserine upon infection. NK-2
was also reported to kill the intracellular parasite Try-
panosoma cruzi, the causative agent of Chagas disease,
leaving the host cell unharmed [15].
Angiotensin II and a related peptide, vanicere-5,
were studied in culture and in mosquitos for their
effects on the development of Plasmodium gallinaceum
sporozoites. When injected into the insect thoraxes,
these peptides reduced infection intensities in the mos-
quito salivary glands by 88% and 76%, respectively.
Although the mechanism of action is not fully under-
stood, the authors proposed that these peptides selec-
tively disrupt the cell membrane of P. gallinaceum, and
showed additionally that preincubation of sporozoites
in vitro with vanicere-5 reduced the infectivity of the
parasites with regard to their vertebrate host [16].
Bombinin H4 is a native antimicrobial peptide of
animal origin with a single l-amino acid to d-amino
acid isomerization in position 2, which improves its
biostability. Bombinin H4 was reported to affect the
viability of both insect and mammalian forms of Leish-
mania by rapidly perturbing their plasma membranes
at micromolar concentrations. The mode of action
involved rapid (within minutes) depolarization and loss
of integrity of the plasma membrane, associated with
rapid bioenergetic collapse [17].
The 13-residue-long temporins show leishmanicidal
activity while maintaining biological functions in
serum. Their lethal mechanism is believed to involve
plasma membrane disruption, on the basis of the
observations that the peptides induce rapid collapse of
the plasma membrane potential, influx of exogenic
molecules, and reduced intracellular ATP levels [18].
Buforin activity against Cryptosporidium parvum was
strongly dependent on the parasite life cycle: the
oocyst was barely affected after 3 h of incubation with
10 lm buforin, whereas the sporozoite’s viability
decreased by almost 100%. The authors speculate that
the sporozoites are susceptible because their cytoplas-
mic membrane is somewhat structurally similar to the
bacterial cytoplasmic membrane [19]. This group also
showed that the moderate activity of buforin can be
enhanced upon combination with azithromycin or
minocycline, > 90% parasite reduction being observed
at the highest concentration tested [20].
A recent study investigated the antiparasitic activi-
ties of different antimicrobial peptides isolated from
aquatic animals. These included penaeidin-3 from the
shrimp Litopenaeus vannamei
, the antilipopolysaccha-
ride factor (ALF) from Penaeus monodon, clavanin A
from the ascidean Styela clava, an analog of magainin
(MSI-94) from the frog Xenopus laevis, tachyplesin I
from the limulid Tachypleus tridentatus, and mytilin A
from the mussel Mytilus edulis. These antimicrobial
peptides were selected because of their previously dem-
onstrated potent effects against bacteria, yeasts, and
filamentous fungi. The antiparasitic activity was eval-
uated against the promastigote form of Leishmania
braziliensis as well as against the epimastigote and
trypomastigote forms of Trypanosoma cruzi. Tachylep-
sin was found to be the most potent peptide in killing
both L. braziliensis and T. cruzi, and was therefore
suggested to be the most suitable candidate for further
investigation as a therapeutic agent [21]. A tachylep-
Antiparasitic peptides A. Mor
6476 FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS
sin-related peptide called gomesin was tested against
asexual, sexual and presporogonic forms of P. falcipa-
rum and P. berghei parasites. When added to culture
of P. berghei mature gametocytes, gomesin inhibited
the exflagellation of male gametes and the formation
of ookinetes. In vivo, the peptide reduced the number
of oocysts of both Plasmodium species in Anophe-
les stephensi mosquitoes [22].
Attacin is an immune effector peptide that has been
shown to inhibit the growth of Gram-negative bacte-
ria. In Glossina morsitans morsitans, which serves as
the sole vector of African trypanosomes, attacin was
implicated in trypanosome resistance and in maintain-
ing parasite numbers at homeostatic levels in infected
individuals [23].
Studies of the histidine-rich antimicrobial peptide
LAH4 resulted in several active derivatives [24]. The
most selective APP was a 26 amino acid analog,
D-HALO-rev, which showed high potency (IC
50
of
0.1 lm) against the human malarial parasite P. falci-
parum, and concentrations toxic against erythrocytes
and fibroblasts at least two orders of magnitude
higher than those needed for its antiplasmodial activ-
ity. The mechanism of the antiplasmodial activity is
unclear; however, important differences in the mem-
brane composition of Plasmodium spp. versus the
host cells are predicted to enhance the activity of
designed antiplasmodial peptides (for a recent review
see [25]). By contrast, another histidine-rich APP,
human histatin-5 (Hst5), seems to target mitochon-
drial ATP synthesis [26]. In the human parasitic pro-
tozoan Leishmania, mitochondrial ATP production is
crucial, as the organism lacks the bioenergetic switch
between glycolysis and oxidative phosphorylation
described in some yeasts. Hst5 displays activity
against both stages of the parasite life cycle, prom-
astigotes and amastigotes (LC
50
values of 7.3 lm and
14.4 lm, respectively). Hst5 was proposed to induce a
lethal effect by causing limited and temporary dam-
age to the plasma membrane of the parasites, as
assessed by electron microscopy, membrane depolar-
ization, and uncontrolled entrance of a vital dye. Fol-
lowing this initial interaction, Hst5 translocates into
the cytoplasm of Leishmania in a nonstereospecific,
receptor-independent manner, accumulates in the
mitochondrion, and produces bioenergetic collapse of
the parasite by decreasing the synthesis of mitochon-
drial ATP.
Tyrothricin, a complex mixture of antibiotic peptides
produced by Bacillus brevis, was reported in 1944 to
have antimalarial activity rivaling that of quinine in
chickens infected with P. gallinaceum. More than
60 years later, the major components of tyrothricin,
cyclic decapeptides collectively known as the tyroci-
dines, were isolated and tested against P. falciparum.
Although the tyrocidines differ from each other by
conservative amino acid substitutions in only three
positions, their parasite inhibitory concentrations
spanned three orders of magnitude (IC
50
of the most
potent compounds ranged between 0.6 and 360 nm).
For comparison, gramicidin S, a structurally analogous
antibiotic peptide tested under the same conditions,
was not as active (IC
50
of 1.3 lm) but exerted anti-
parasitic activity by rapid lysis of infected erythro-
cytes. Like those of previously described antimicrobial
peptides, tyrocidine activities correlated strictly with
increased apparent hydrophobicity and reduced total
side chain surface area due to the presence of ornithine
and phenylalanine in key positions. Unlike antiplasmo-
dial activity, however, mammalian cell toxicities of the
respective peptides were considerably less variable,
ranging only from 2.6 to 28 lm [27].
Various native members of the frog-derived derm-
aseptin family exhibit potent antiparasitic properties
against both Leishmania [28] and P. falciparum [29].
Synthetic derivatives of dermaseptin S4, such as the
28-mer K
4
K
20
-S4 or its short analog K
4
S4(1–13), dis-
played enhanced activity towards human erythrocytes
infected with P. falciparum, killing the parasite through
lysis of the host cells [30]. Both derivatives were more
efficient in inhibiting parasite growth at the mature
trophozoite stage than at the younger ring stage. This
fact supports the view that the antiplasmodial effect is
essentially derived from lysing the host cell membrane;
that is, because the host cell membrane evolves with
parasite maturation, trophozoites are expected to be
more sensitive than rings, as observed [30]. Various
conjugation derivatives of K
4
S4(1–13) were assayed
with the aim of avoiding lysis of host cells. These
derivatives have established that increased hydropho-
bicity at the N-terminus invariably results in an ampli-
fied antiplasmodial effect, irrespective of the linearity
or bulkiness of the additive. However, increased
hydrophobicity was also generally associated with
increased hemolysis and lack of discrimination between
infected and noninfected erythrocytes [31]. By contrast,
aminoacyl counterparts were generally more selective
[32]. Thus, as compared with the parent peptide, the
aminoheptanoylated version displayed both increased
antiparasitic efficiency and reduced hemolysis, includ-
ing against infected cells. Presumably, by selectively
dissipating the parasite plasma membrane potential
and causing depletion of intraparasite potassium, this
derivative exerted more than 50% growth inhibition at
peptide concentrations that did not cause detectable
hemolysis. Hence, unlike the parent peptide, the
A. Mor Antiparasitic peptides
FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS 6477
aminoheptanoylated derivative was not stage-selective,
being equally inhibitory for both the ring and tropho-
zoite stages. Additional new members of the derma-
septin family also act as APPs, displaying potent
activity against Leshmania (e.g. dermaseptins O1 and
H3) and against T. cruzi (e.g. dermaseptins D11 and
D12) [25,38,39].
Chemical mimicry of APPs
In addition to the naturally occurring APPs and their
de novo designed synthetic derivatives, recent studies
suggest that potent antiparasitic properties can be
generated from HDP-mimetic compounds designed to
mimic the structure and ⁄ or function of the native
peptides [33,34]. The potential therapeutic use of anti-
parasitic HDPs is likely to be significantly limited by
several major obstacles pertaining to less than ideal
properties, including relative toxicity and bioavail-
ability issues as well as a relatively high production
cost. By reproducing the critical biophysical character-
istics of HDPs, peptidomimetics might better address
these issues while endowing resistance to degradation
enzymes.
Recently developed HDP mimetics, termed oligo-
acyl-lysyls (OAKs), were based on a linear sequence of
alternating aminoacyl chains and cationic amino acids
so as to mimic the prototypical sequence of linear
HDPs. Like potent HDPs, OAKs can display rapid,
nonhemolytic, broad-spectrum microbicidal properties
both in vitro and in vivo. Various OAK sequences were
shown to inhibit the growth of different plasmodial
strains (IC
50
range 0.08–0.14 lm). Further investiga-
tions performed with a representative OAK revealed
that the ring and trophozoite stages of the parasite
developmental cycle were equally sensitive to this com-
pound, unlike the case with the parent dermaseptin
peptides. A shortcoming of the OAK was the need for
long incubation times in order for it to exert its full
effect [35]. Nevertheless, certain OAKs displayed
highly selective antiparasitic activity, the ratio of LC
50
(hemolysis) to IC
50
(parasite growth inhibition) being
> 10 000 for the most selective OAK, composed of
only three acyl-lysyl subunits (Table 1). These results
indicated that the OAK did not exert its antimalarial
action by lysis of infected erythrocytes, as was the case
with the parent dermaseptins, and pointed to the
potential of the OAK system to generate simple, highly
selective and low-cost compounds that might be useful
in fighting malaria. Note that, although the OAKs rep-
resent, so far, the only HDP mimetic system able to
generate antiparasitic compounds, it is anticipated that
future studies of various existing and ⁄ or new future
mimic strategies will reveal interesting antiparasitic
properties. Because of their chemical robustness, such
compounds are likely to overcome various drawbacks
of conventional antimicrobial peptides, including sus-
ceptibility to proteases, and might therefore be both
useful investigation tools and new, promising candi-
dates for therapeutic developments.
Possible mechanisms for APPs
Attempts to understand the mechanism(s) underlying
the observed antiplasmodial activity suggest that the
activities of distinct APPs obey many of the rules
governing their ability to disrupt bacterial membranes
(discussed extensively elsewhere in this issue). This type
of interaction appears to be acutely influenced by the
respective charges and amphipathies of the reactants.
In fact, a comparison between the peptides’ ability to
inhibit the growth of malaria parasites and that of
bacteria demonstrates a remarkable parallelism in the
way that each modification affects both activities, as
assessed with numerous substitution ⁄ truncation deriv-
atives [28–31]. Additional support for this view is
provided by the results obtained with experiments on
kinetics and the dissipation of the membrane potential,
as well as from the fact that activity is rapid and is
independent of a chiral center [32]. Therefore, a variety
of experimental evidence suggests that the mode of
antiplasmodial activity of some antimicrobial peptides
might be based on selective membrane disruption,
despite the fact that the parasite’s membrane is well
hidden within its host cell. As shown in Fig. 1, it is
speculated that, owing to differences in membrane
composition [25,40], APPs have a higher affinity for
the membranes of infected erythrocytes than for those
of normal erythrocytes (hence the often observed
greater extent of hemolysis of infected cells), but must
have a still higher affinity for the parasite’s membrane.
The observed labeling of intracellular parasites in non-
lysed, infected erythrocytes (i.e. under nonhemolytic
conditions) supports this view (Fig. 1), although the
apparent differential distribution of rhodaminated
peptides may also be due to an experimental artefact:
the fluorescence may be collisionally quenched by
hemoglobin, which is present only in the host cell com-
partment. Nevertheless, colocalization evidence exists
(although unpublished) suggesting that, in macro-
phage-infected amastigotes, the antileishmanial activity
also might proceed by direct interaction of dermasep-
tin with the intracellular parasite (Fig. 2). Thus, when
dermaseptin, for instance, binds the membrane of a
hosting erythrocyte, the peptide is somehow able to
transfer to the parasitic membrane in an affinity-driven
Antiparasitic peptides A. Mor
6478 FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS
Fig. 1. Proposed model for APP interaction with infected erythrocytes. The left panel shows phase-contrast and fluorescence confocal
microscope images providing evidence for the direct interaction of the rhodamine-labeled dermaseptin derivative, aminohexyl-K
4
-S4(1–13),
with unfixed intraerythrocyte P. falciparum trophozoites. The right panel is a cartoon illustration of two hypothetical modes of action. The
upper drawing shows the initial adhesion of the lipophilic APP (blue) to the erythrocyte membrane. Subsequently, hemolytic APPs can
assemble to disrupt that membrane (right drawing). The resulting hemoglobin leakage will lead to parasite starvation. Nonhemolytic APPs
(left drawing) can undergo an affinity-driven transfer (through lateral diffusion?) from the erythrocyte to the parasite membranes. The lower
drawings represent potential APP fates once they have reached the parasite’s membrane: (1) superficial carpet-like interactions can modify
the membrane properties (e.g. charge and fluidity of lipids and protein components) and interfere with their proper normal functions [40]; (2)
APP internalization and interruption of vital biological processes (e.g. DNA functions) [34]; (3) APPs can also disrupt the parasite’s plasma
membrane (similarly to the hemolytic process) as evidenced by the APP’s ability to dissipate the parasite plasma membrane potential and
cause depletion of intraparasite potassium under nonhemolytic conditions [32].
A
B
C
D
Fig. 2. Proposed model for APP interactions with infected macrophages. The left panel shows the experimental colocalization of APP and
parasites in a human macrophage infected with L. major amastigotes, as visualized by fluorescence confocal microscopy: (A) rhodaminated
dermaseptin applied to unfixed cells; (B) fluoresceinated antiparasite antibody applied after cell fixation; (C) merged image of (A) and (B); (D)
phase-contrast image of the infected macrophage shown in (A), (B) and (C). The right panel is a cartoon illustration of two hypothetical
routes leading to the colocalization of dermaseptin and parasite in an infected macrophage. In the upper drawings, the cartoon shows that
cationic APPs could reach the cytoplasm by diffusion, exploiting the negative-inside transmembrane potential or via vesicle-like internalization
followed by fusion with the parasitophorous vacuole. The subsequent events that might follow are described in the lower drawings of
Fig. 1.
A. Mor Antiparasitic peptides
FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS 6479
manner and to exert a membrane-lytic activity upon
the pathogen. Namely, such a ‘transfer’ could physi-
cally occur through the new permeability pathways
that are induced by the parasite in the membrane of
the host erythrocyte. As the parasite is completely
engulfed within a parasitophorous vacuole membrane,
solutes which leave or enter the parasite must therefore
traverse three membranes: that of the host erythrocyte,
the parasitophorous vacuole membrane, and the para-
site membrane. Experimental evidence for the ability
of APPs to gain access to the parasite in malaria-
infected cells, obtained using confocal microscopy
analysis of labeled dermaseptins, is shown in Fig. 1
[32]. The data showed that, in infected cells, the
labeled peptide crossed the erythrocyte plasma mem-
brane and concentrated in internal compartments,
although it is presently unclear what was the peptide’s
ultimate target, e.g. the parasite membrane, the mito-
chondrion, or nucleic acids. Antimicrobial peptides are
known to target each and ⁄ or all of the above [34].
Conclusions
Although the mechanism of action of most antimicro-
bial peptides is far from being fully understood, the
vast majority are now believed to act by one or even a
combination of different nonspecific mechanisms that
can target not only the cell membrane integrity but
also extracellular and intracellular processes (reviewed
by Pierre Nicolas in [41]; this issue). Such a multitarget
mode of action is in good agreement with the observed
large spectrum of sensitive microorganisms, and signifi-
cantly reduces the likelihood of emergence of efficient
resistance mechanisms. Thus, even though the antipar-
asitic properties have not been investigated thoroughly,
as yet, an increasing number of convincing studies
seem to support the view that the antiparasitic activity
of antimicrobial peptides also emanates from interac-
tions with multiple targets. Most remarkably, however,
at least a few peptides exhibit very high potency (IC
50
values in the nanomolar range) and a selectivity factor
of several orders of magnitude. These selective com-
pounds appear to be endowed with the formidable
ability to cross a number of membrane systems before
specifically disrupting a target(s) in the intracellular
parasite. Although differences in membrane composi-
tion are likely to contribute to this selectivity, the
molecular basis for these observations remains largely
ill-understood. Nonetheless, these studies strongly sug-
gest that the physicochemical attributes resulting from
the molecular structure of antimicrobial peptides can
be useful in engineering selective and efficient antipara-
sitic therapeutic drugs.
Acknowledgement
This research was supported by the Israel Science
Foundation (grant 283 ⁄ 08).
References
1 Korenromp EL (2004) Malaria Incidence Estimates at
Country Level for the Year 2004 – Proposed Estimates
and Draft Report. Roll Back Malaria, World Health
Organization, Geneva.
2 Bruce-Chwatt LJ (1982) Chemoprophylaxis of malaria
in Africa: the spent ‘magic bullet’. BMJ 285, 674–676.
3 Krishna S, Woodrow CJ, Staines HM, Haynes RK &
Mercereau-Puijalon O (2006) Re-evaluation of how
artemisinins work in light of emerging evidence of
in vitro resistance. Trends Mol Med 12, 200–205.
4 World Health Organization (2005) World Malaria
Report 2005. WHO, Geneva. http://www.rbm.who.int/
wmr2005.
5 Vizioli J & Salzet M (2002) Antimicrobial peptides
versus parasitic infections. Trends Parasitol 18, 475–
476.
6 Gwadz RW, Kaslow D, Lee JY, Maloy WL, Zasloff M
& Miller LH (1989) Effects of magainins and cecropins
on the sporogonic development of malaria parasites in
mosquitoes. Infect Immun 57, 2628–2633.
7 Boman HG, Wade D, Boman IA, Wahlin B &
Merrifield RB (1989) Antibacterial and antimalarial
properties of peptides that are cecropin–melittin
hybrids. FEBS Lett 259, 103–106.
8 Tian C, Gao B, Rodriguez MC, Lanz-Mendoza H, Ma
B & Zhu S (2008) Gene expression, antiparasitic activ-
ity, and functional evolution of the drosomycin family.
Mol Immunol 45, 3909–3916.
9 Conde R, Zamudio FZ, Rodriguez MH & Possani LD
(2000) Scorpine, an anti-malaria and anti-bacterial
agent purified from scorpion venom. FEBS Lett 471,
165–168.
10 Carballar-Lejarazu R, Rodriguez MH, de la Cruz Her-
nandez-Hernandez F, Ramos-Castaneda J, Possani LD,
Zurita-Ortega M, Reynaud-Garza E, Hernandez-Rivas
R, Loukeris T, Lycett G et al. (2008) Recombinant
scorpine: a multifunctional antimicrobial peptide with
activity against different pathogens. Cell Mol Life Sci
65, 3081–3092.
11 Kulkarni MM, McMaster WR, Kamysz E, Kamysz W,
Engman DM & McGwire BS (2006) The major surface-
metalloprotease of the parasitic protozoan, Leishmania,
protects against antimicrobial peptide-induced apoptotic
killing. Mol Microbiol 62, 1484–1497.
12 Tsuji N, Battsetseg B, Boldbaatar D, Miyoshi T, Xuan
X, Oliver JH Jr & Fujisaki K (2007) Babesial vector
tick defensin against Babesia sp. parasites. Infect Immun
75, 3633–3640.
Antiparasitic peptides A. Mor
6480 FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS
13 Leitch GJ & Ceballos C (2009) A role for antimicrobial
peptides in intestinal microsporidiosis. Parasitology 136,
175–181.
14 Gelhaus C, Jacobs T, Andra J & Leippe M (2008) The
antimicrobial peptide NK-2, the core region of mamma-
lian NK-lysin, kills intraerythrocytic Plasmodium falci-
parum. Antimicrob Agents Chemother 52, 1713–1720.
15 Jacobs T, Bruhn H, Gaworski I, Fleischer B & Leippe
M (2003) NK-lysin and its shortened analog NK-2
exhibit potent activities against Trypanosoma cruzi.
Antimicrob Agents Chemother 47, 607–613.
16 Maciel C, de Oliveira VX Jr, Fazio MA, Nacif-Pimenta
R, Miranda A, Pimenta PF & Capurro ML (2008)
Anti-plasmodium activity of angiotensin II and related
synthetic peptides. PLoS ONE 3, e3296.
17 Mangoni ML, Papo N, Saugar JM, Barra D, Shai Y,
Simmaco M & Rivas L (2006) Effect of natural L- to
D-amino acid conversion on the organization, mem-
brane binding, and biological function of the antimicro-
bial peptides bombinins H. Biochemistry 45, 4266–4276.
18 Mangoni ML, Saugar JM, Dellisanti M, Barra D,
Simmaco M & Rivas L (2005) Temporins, small anti-
microbial peptides with leishmanicidal activity. J Biol
Chem 280, 984–990.
19 Giacometti A, Cirioni O, Del Prete MS, Barchiesi F &
Scalise G (2000) Short-term exposure to membrane-
active antibiotics inhibits Cryptosporidium parvum
infection in cell culture. Antimicrob Agents Chemother
44, 3473–3475.
20 Giacometti A, Cirioni O, Del Prete MS, Barchiesi F,
Fineo A & Scalise G (2001) Activity of buforin II alone
and in combination with azithromycin and minocycline
against Cryptosporidium parvum in cell culture. J Anti-
microb Chemother 47, 97–99.
21 Lofgren SE, Miletti LC, Steindel M, Bachere E & Barr-
acco MA (2008) Trypanocidal and leishmanicidal activi-
ties of different antimicrobial peptides (AMPs) isolated
from aquatic animals. Exp Parasitol 118, 197–202.
22 Moreira CK, Rodrigues FG, Ghosh A, Varotti FP,
Miranda A, Daffre S, Jacobs-Lorena M & Moreira LA
(2007) Effect of the antimicrobial peptide gomesin
against different life stages of Plasmodium spp. Exp
Parasitol 116, 346–353.
23 Hu Y & Aksoy S (2005) An antimicrobial peptide with
trypanocidal activity characterized from Glossina morsi-
tans morsitans. Insect Biochem Mol Biol 35, 105–115.
24 Mason AJ, Moussaoui W, Abdelrahman T, Boukhari
A, Bertani P, Marquette A, Shooshtarizaheh P, Moulay
G, Boehm N, Guerold B et al. (2009) Structural deter-
minants of antimicrobial and antiplasmodial activity
and selectivity in histidine-rich amphipathic cationic
peptides. J Biol Chem 284, 119–133.
25 Rivas L, Luque-Ortega JR & Andreu D (2008) Amphib-
ian antimicrobial peptides and Protozoa: lessons from
parasites. Biochim Biophys Acta 1788, 1570–1581.
26 Luque-Ortega JR, Vant HW, Veerman EC, Saugar JM
& Rivas L (2008) Human antimicrobial peptide
histatin 5 is a cell-penetrating peptide targeting mito-
chondrial ATP synthesis in Leishmania. FASEB J 22,
1817–1828.
27 Rautenbach M, Vlok NM, Stander M & Hoppe HC
(2007) Inhibition of malaria parasite blood stages by
tyrocidines, membrane-active cyclic peptide antibiotics
from Bacillus brevis. Biochim Biophys Acta 1768, 1488–
1497.
28 Hernandez C, Mor A, Dagger F, Nicolas P, Hernandez
A, Benedetti EL & Dunia I (1992) Functional and
structural damage in Leishmania mexicana exposed to
the cationic peptide dermaseptin. Eur J Cell Biol 59,
414–424.
29 Ghosh JK, Shaool D, Guillaud P, Ciceron L, Mazier
D, Kustanovich I, Shai Y & Mor A (1997) Selective
cytotoxicity of dermaseptin S3 toward intraerythrocytic
Plasmodium falciparum and the underlying molecular
basis. J Biol Chem 272, 31609–31616.
30 Krugliak M, Feder R, Zolotarev VY, Gaidukov L,
Dagan A, Ginsburg H & Mor A (2000) Antimalarial
activities of dermaseptin S4 derivatives. Antimicrob
Agents Chemother 44, 2442–2451.
31 Dagan A, Efron L, Gaidukov L, Mor A & Ginsburg H
(2002) In vitro antiplasmodium effects of dermasep-
tin S4 derivatives. Antimicrob Agents Chemother 46,
1059–1066.
32 Efron L, Dagan A, Gaidukov L, Ginsburg H & Mor A
(2002) Direct interaction of dermaseptin S4 aminohep-
tanoyl derivative with intraerythrocytic malaria parasite
leading to increased specific antiparasitic activity in cul-
ture. J Biol Chem 277, 24067–24072.
33 Scott RW, DeGrado WF & Tew GN (2008) De novo
designed synthetic mimics of antimicrobial peptides.
Curr Opin Biotechnol 19, 620–627.
34 Rotem S & Mor A (2008) Antimicrobial peptide mimics
for improved therapeutic properties. Biochim Biophys
Acta 1788, 1582–1592.
35 Radzishevsky I, Krugliak M, Ginsburg H & Mor A
(2007) Antiplasmodial activity of lauryl-lysine oligo-
mers. Antimicrob Agents Chemother 51, 1753–1759.
36 Soravia E, Martini G & Zasloff M (1988) Antimicrobial
properties of peptides from Xenopus granular gland
secretions. FEBS Lett 228, 337–340.
37 Madison MN, Kleshchenko YY, Nde PN, Simmons
KJ, Lima MF & Villalta F (2007) Human defensin
alpha-1 causes Trypanosoma cruzi membrane pore
formation and induces DNA fragmentation, which
leads to trypanosome destruction. Infect Immun 75,
4780–4791.
38 Brand GD, Leite JR, Silva LP, Albuquerque S, Prates
MV, Azevedo RB, Carregaro V, Silva JS, Sa
´
VC, Bran-
da
˜
oRAet al. (2002) Dermaseptins from Phyllomedusa
oreades and Phyllomedusa distincta. Anti-Trypanosoma
A. Mor Antiparasitic peptides
FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS 6481
cruzi activity without cytotoxicity to mammalian cells.
J Biol Chem 277, 49332–49340.
39 Brand GD, Leite JR, de Sa
´
Mandel SM, Mesquita DA,
Silva LP, Prates MV, Barbosa EA, Vinecky F, Martins
GR, Galasso JH et al. (2006) Novel dermaseptins from
Phyllomedusa hypochondrialis. Biochem Biophys Res
Commun 347, 739–746.
40 Epand RM & Epand RF (2009) Lipid domains in bac-
terial membranes and the action of antimicrobial
agents. Biochim Biophys Acta 1788, 289–294.
41 Nicolas P (2009) Multifunctionalhostdefense peptides:
Intracellular-targeting antimicrobial peptides. FEBS J
276, doi:10.1111/j.1742-4658.2009.07359.x.
Antiparasitic peptides A. Mor
6482 FEBS Journal 276 (2009) 6474–6482 ª 2009 The Author Journal compilation ª 2009 FEBS
. MINIREVIEW Multifunctional host defense peptides: antiparasitic activities Amram Mor Department of Biotechnology & Food Engineering,. 289–294. 41 Nicolas P (2009) Multifunctional host defense peptides: Intracellular-targeting antimicrobial peptides. FEBS J 276, doi:10.1111/j.1742-4658.2009.07359.x. Antiparasitic peptides A. Mor 6482. and antifungal properties of host defense peptides (HDPs) and their synthetic mimics, much less is known of their activities against parasites. A variety of in vitro and in vivo antiparasitic assays