Advances and challenges towards a vaccine against Chagas disease.

Abstract

Chagas disease is major public health problem, affecting nearly 10 million people, characterized by cardiac alterations leading to congestive heart failure and death of 20-40% of the patients infected with Trypanosoma cruzi, the protozoan parasite responsible for the disease. A vaccine would be key to improve disease control and we review here the recent advances and challenges of a T. cruzi vaccine. There is a growing consensus that a protective immune response requires the activation of a Th1 immune profile, with the stimulation of CD8 (+) T cells. Several vacines types, including recombinant proteins, DNA and viral vectors, as well as heterologous prime-boost combinations, have been found immunogenic and protective in mouse models, providing proof-of-concept data on the feasibility of a preventive or therapeutic vaccine to control a T. cruzi infection. However, several challenges such as better end-points, safety issues and trial design need to be addressed for further vaccine development to proceed.

Full text

Page 1

©2011 Landes Bioscience.
Do not distribute.
www.landesbioscience.com Human Vaccines 1
Human Vaccines 7:11, 1-8; November 2011; © 2011 Landes Bioscience
Advances and challenges toward a vaccine
against chagas disease
SpeciaL FocuS ReView: NegLected VacciNeS—deVeLopiNg woRLd
SpeciaL FocuS ReView: NegLected VacciNeS—deVeLopiNg woRLd
Introduction
Chagas disease or American Trypanosomiasis is major public
health problem, affecting nearly 10 million people, mostly in
Latin America where it is endemic, but international migrations
are making the disease increasingly frequent in non-endemic
countries as well. It is one of the most neglected tropical diseases,
frequently associated with poverty and margination in rural
areas, although it is also expanding in richer suburban and urban
areas in many countries. The protozoan parasite Trypanosoma
cruzi is the causative agent of Chagas disease, and it is trans-
mitted mainly by hematophagous vectors from the triatominae
family, commonly known as kissing bugs. Secondary routes of
transmission include blood transfusion, organ transplant and
materno-fetal transmission, and oral outbreaks have also been
described.
The disease initiates by an acute phase lasting a few weeks
and characterized by an elevated parasitemia associated with
fever, headache, nausea, that is rarely lethal. This phase is fol-
lowed by a chronic phase, which remains asymptomatic for many
years. The parasites are then difficult to observe in the blood,
*Correspondence to: Eric Dumonteil; Email: oliver@uady.mx
Submitted: 06/19/11; Revised: 07/17/11; Accepted: 07/22/11
DOI:
chagas disease is major public health problem, affecting nearly
10 million people, characterized by cardiac alterations leading
to congestive heart failure and death of 20–40% of the patients
infected with Trypanosoma cruzi, the protozoan parasite
responsible for the disease. a vaccine would be key to improve
disease control and we review here the recent advances and
challenges of a T. cruzi vaccine. there is a growing consensus
that a protective immune response requires the activation of a
th1 immune profile, with the stimulation of cd8+ t cells. Several
vacines types, including recombinant proteins, dNa and viral
vectors, as well as heterologous prime-boost combinations,
have been found immunogenic and protective in mouse
models, providing proof-of-concept data on the feasibility of a
preventive or therapeutic vaccine to control a T. cruzi infection.
However, several challenges such as better end-points, safety
issues and trial design need to be addressed for further vaccine
development to proceed.
israel Quijano-Hernandez1,2 and eric dumonteil1,3,*
1Laboratorio de parasitología; centro de investigaciones Regionales ‘dr. Hideyo Noguchi; universidad autónoma de Yucatán; Merida; 2Hospital Veterinario para pequeñas
especies de la Facultad de Medicina Veterinaria y Zootecnia de la universidad autónoma del estado de México; toluca, México; 3department of tropical Medicine; School of
public Health and tropical Medicine; tulane university; New orleans, La uSa
Key words: American trypanosomiasis, Trypanosoma cruzi, vaccine, animal model, pre-clinical development
This manuscript has been published online, prior to printing. Once the issue is complete and page numbers have been assigned, the citation will change accordingly.
and diagnosis relies principally on serologic tests. The majority of
patients remain in this indeterminate stage for life, while 20–40%
start presenting a symptomatic chronic phase characterized by a
progressive and debilitating Chagasic cardiomyopathy and some-
times megasyndromes. Arrhythmias of increasing severity lead to
congestive cardiac failure and death of the patients.1
Drug treatments of infected patients are based on Nifurtimox
and Benznidazol since the 1960s, and while their efficacy is fairly
well established during the acute phase of infection and in chil-
dren, their usefulness in chronic phase and adult patients is still
debated and a large clinical trial is currently underway to clarify
this point.2,3 Both drugs require prolonged treatments and can
have severe adverse effects.4,5 Patients also require palliative care
for the cardiomyopathy, ranging from drug therapy to pacemak-
ers and cardiac transplants.6 Thus treatment can cost up to over
1,000 USD per year and per patient.7
Prevention currently only relies on vector control, which has
considerably reduced disease incidence and prevalence in most
Latin America.8 Nonetheless, a number of triatomine species have
proven difficult to control and the most likely scenario is that low
level vectorial transmission to humans will be mantained in may
regions and countries in spite of vector control interventions.9
In this context, a vaccine would be key to improve the con-
trol of Chagas disease, and a recent study highlighted the large
cost-effectiveness that a preventive vaccine for humans would
have in a variety of scenarios. Thus, a vaccine would be cost-
effective even where transmission to humans and prevalence of
infection are low (i.e., after vector control or in regions of low
endemicity), and even for a vaccine of moderate protective effi-
cacy.10 Alternatively, a therapeutic vaccine administered alone or
in combination with current drug treatment may greatly improve
the prognosis for Chagasic patients by increasing treatment effi-
cacy, reducing its duration and cost, or at least delaying disease
progression to advanced stages and heart failure. Such a thera-
peutic vaccine is also likely to be very cost effective. In addition,
in some regions, domestic animals such as dogs, have been found
to play an important role as reservoir of the parasite and its trans-
mission to human.11-14 Control of infection in these animals has
thus been proposed as an important complementary strategy to
vector control, for example using insecticide-impregnated col-
lars.15 However, a veterinary vaccine, either preventive or thera-
peutic, would thus also greatly contribute to Chagas disease
control by reducing infectiousness of the main domestic reservoir

Page 2

©2011 Landes Bioscience.
Do not distribute.
by dendritic cells allowing for its persistence, while in a model
of dendritic cell-based immunization, IL-10-deficient dendritic
cells pulsed with T. cruzi antigens conferred protection against
infection to recipient mice.25 Specific parasite proteins such as
cruzipain and Tc52 in can directly increase IL-10 expression,
leading to a non-protective immune response.26 Thus, aim-
ing at both early killing of parasites and neutralizing suppres-
sive mechanisms may be necessary for effective therapies and
vaccines.
IFNγ plays a key role during T. cruzi infection, increasing the
production of nitric oxide by macrophages, which inhibits the
development of the intracellular form of T. cruzi.27 CD4+ Th1
lymphocytes that produce the cytokines interleukin-2 (IL-2) and
IFNγ and can stimulate the expansion of cytotoxic CD8+ T lym-
phocytes, which appear central for systemic protection against
T. cruzi infection.28,29 Recent studies have confirmed that CD8+
T cells act both indirectly by secreting IFNγ to activate macro-
phages and directly through the production of perforin and their
concomitant cytotoxic activity.30 IFNγ producing CD4+ and
CD8+ T cells also play a key role in the liver for the clearance of
blood trypomastigotes.31 These cells are also stimulated follow-
ing oral infection, which is supposed to be a common route of
infection for several animal reservoir species.32 Thus, there is now
a growing consensus that a protective immune response against
T. cruzi requires the activation of a Th1 immune profile, with the
stimulation of CD8+ T cells, while antibodies may play a rather
secondary role.
2 Human Vaccines Volume 7 issue 11
Current Status of Vaccine Research
A very wide range of vaccine formulations have been evaluated
over the years, ranging from whole parasites, to purified or recom-
binant proteins, viral vectors and DNA vaccines (Table 1). Initial
attempts at using killed T. cruzi parasites resulted in modest but
encouraging protection against infection,33 possibly because of
a rather low immunogenicity. Live attenuated parasites on the
other hand are much more immunogenic and early studies indi-
cated that they were able to confer significant protection against
infection.34 Similarly, immunization with live T. rangeli, a related
parasite species non-pathogenic in humans, can also provide
some cross-reactive immunity and partial protection, possibly
due to homologous antigens.35-37 However, these approaches raise
the issues commonly associated with live vaccines, i.e., safety and
the challenges associated with their large-scale production and
distribution. Nonetheless, there is a renewed interest in a live
attenuated vaccine approach and recent studies have described
the generation of T. cruzi mutants for specific genes such as LYT1
or ECH1 and 2, which can provide good protection against infec-
tion in mice, including through oral administration.32,38
Early attempts at immunizing mice with more defined prep-
arations were based on proteins purified from parasite culture,
including cruzipain, the paraflagellar rod protein or trypomasti-
gote excretory-secretory antigens.39-41 As expected, the outcome
of such vaccines is critically dependent on the formulation used,
and the orientation of the immune response that is induced. For
example, immunization with cruzipain administered with alum
or Freund adjuvants, which bias the immune response toward
a Th2 type and high antibody levels, is not protective. On the
other hand, the same antigen administered with CpG oligonucle-
otides induces a Th1 type immune response with elevated IFNγ
production, which protects mice against infection by reducing
parasitemia and increasing survival.42 More recent studies are
based on the use of recombinant proteins, which have now been
extensively evaluated. Some of the most promising antigens tested
include cruzipain, GP82, and several proteins from the trans-
sialidase superfamily such as ASP-2 and TS, and all can provide
significant protection in a variety of mouse models when admin-
istered with CpG oligonucleotides as adjuvants (Table 1). This
again, highlights the need to induce a strong cellular immune
response with CD8+ cell activation and cytotoxic activity for the
control of T. cruzi infection, rather than an antibody response.
Accordingly, several of these antigens have been formulated
as recombinant viral vaccine vectors as well as DNA vaccines
since these formulations may be more effective than recombinant
proteins at inducing a Th1 type immune response with cyto-
toxic CD8+ T cells.43,44 Sendai, vaccinia and adenovirus vectors
are among those that have been evaluated, expressing several of
the antigens mentioned above, including ASP-2 and TS proteins
from the trans-sialidase superfamily (Table 1). As expected such
vaccines were able to induce strong IFNγ responses as well as
the activation of CD8+ cytotoxic T cells, which led to a strong
protection against infection as indicated by a decreased parasit-
emia and increased survival of mice. Such protection was often
better than that achieved by the corresponding recombinant
of the parasite, as well as improving the care of companion ani-
mals that are increasingly been detected as infected by T. cruzi.16
We review here some of the recent advances and challenges in T.
cruzi vaccine development, and an analysis of earlier work can be
found in previous reviews in reference 17–19.
Correlates for Immune Protection against T. cruzi
A good understanding of the interactions between T. cruzi and
the immune system is key to the rational design and improve-
ment of vaccines, and important advances have been made in
recent years. However, it is important to recall that T. cruzi
vaccine research has been hindered by the fear of exacerbating
myocardial lesions through an activation of a severe autoimmune
response, based on the hypothesis that Chagas disease was actu-
ally an autoimmune disease.20,21 Nonetheless, parasite persistence
rather than an imbalanced immune response, has been shown to
be associated with disease progression,22 strengthening the ratio-
nale for a vaccine (see also below).
Recognition of T. cruzi by the immune system relies on the
activation of the innate and adaptative immunity. Recognition
of pathogen-associated molecular patterns (PAMP) by Toll-
like receptors (TLR) such as TLR-2 and TLR-9, lead to the
activation of T and B cells, making TLRs an important link
between innate and adaptative immunity. The absence of TLR
signaling reduces the activation of CD4+ cells against T. cruzi,
while CD8+ responses may be maintained.23 Indeed, T. cruzi is
able to downregulate the immune system through the release of
several molecules.23,24 T. cruzi can promote IL-10 production

Page 3

©2011 Landes Bioscience.
Do not distribute.
iFNγ, cd8+,
cytolytic
activity
www.landesbioscience.com Human Vaccines 3
expressing a single dominant CD8+ epitope,46 which may provide
additional alternatives for immunization. Indeed, immunization
with a vaccinia and adenovirus expressing this epitope was previ-
ously found to be sufficient to induce protection, highlighting the
central role of this T cell population in parasite control.47 In fact,
highly immunodominant CD8+ epitopes have been identified in
the trans-sialidase protein family, resulting in a restricted immune
response during T. cruzi infection, as observed in both mice and
humans.48,49 This explains why most vaccine studies have focused
protein, strengthening the idea that vaccine type is key to achieve
the desired immune response. Furthermore, heterologous prime-
boost immunization strategies, based on a combination of DNA
immunization followed by recombinant protein or viral vectors,
have also been evaluated and may further enhance the protec-
tive immunity induced. However, such complicated immuniza-
tion protocols may be difficult to translate into a field vaccine.
New recombinant virus vectors have also recently be described,
such as Yellow fever virus expressing ASP-2 45 and influenza virus
Table 1. preventive vaccines against T. cruzi evaluated in mice
FormulationAntigens
Immune
response
ParasitemiaEnd points
Heart
Histopathology
SurvivalReference
whole organism
Live attenuated
Trypanosoma cruzi
NRdecreased decreased*NRNR 32
Live Trypanosoma
rangeli
NR decreaseddecreasedNo changeincreased37
Recombinant
protein
raSp-2 + alum or cpg
odN
increased iFNγ
decreasedNRNR increased94, 95
rtS (trans-sialidase) +
cpg odN
iga, igg and
iFNγ
NRdecreasedNRincreased 96, 97
rcruzipain + cpg odN
Humoral
and cellular
response, iFNγ
and lytic
activity
decreased NR
decreased inflam-
mation**
NR 98
rgp82 + cpg odN
iFNγ
NRNR NR
Variable
according
to challenge
route
99
Viral vector
adenovirus or vaccinia
expressing tSSa cd8+
epitope
iFNγ
decreased NRNRincreased 47
adenovirus expressing
tS and aSp-2
antibodies
and cytolytic
activity
decreased NRNRincreased100
Sendai virus express-
ing aSp-2
iFNγ and cd8+
decreased NRNRincreased 101
dNa
tSa-1, aSp-1 and
aSp-2 with iL-12 and
gM-cSF
antibodies
and cytolytic
activity
decreasedNR
decreased inflam-
mation
increased 102
aSp-2 or uB-aSp-2
decreased
NRNR
increased
94, 103
tS + iL15
iFNγ and cd8+
NR NRNRincreased104
tSa-1 and tc24NR decreasedNR
decreased inflam-
mation
NR60
Heterologous
prime-boost
tcVac2 (tcg1, tcg2,
tcg4) + iL-12 +
gM-cSF dNa and
recombinant protein
boost + saponin
Humoral
and cellular
response, iFNγ
and cd8+
decreaseddecreasedincreasedNR 53
dNa + adenovirus
expressing tS and
aSp-2 clone 9 (prime-
Boost)
iFNγ, cd8+,
cytolytic
activity
decreased NRNRincreased105
NR: Not reported. *parasite burden was actually measured in skeletal muscle rather than heart. **No alterations were observed in the heart, but inflam-
mation was reduced in skeletal muscle.

Page 4

©2011 Landes Bioscience.
Do not distribute.
from acute parasitemia and chronic cardiac tissue damage.52,53
Alternatively, immunization with a trypomastigote cDNA
library and the sequential selection of protective pools has also
been used to identify a series of novel antigens able to confer
increased survival following infectious challenge.54 Similarly, a
novel family of surface proteins has been recently identified that
seems to be specific for T. cruzi.55 Taken together, these stud-
ies may thus lead to some promising antigens with important
potential as novel vaccine candidates.
While the studies mentioned above support the feasibility of
a preventive vaccine against T. cruzi, it is also noteworthy that
therapeutic vaccines can control an ongoing T. cruzi infection
(Table 2). Indeed, the administration of DNA vaccines encod-
ing a trans-sialidase antigen (TSA-1) or the secreted antigen
Tc2456 following a T. cruzi infection was found able to reduce the
parasitemia and cardiac inflammatory reaction, while increas-
ing survival of treated mice.57 Similarly, the immunotherapy
with recombinant SA85-1.1 from the trans-sialidase superfamily,
allowed to reduce parasitemia and cardiac tissue damage.58 It is
also important to stress that no exacerbation of disease has been
observed in these studies, in spite of the significant activation of
the immune response following vaccine administration. Indeed,
in agreement with the data presented above on preventive vac-
cine, the efficacy of such therapeutic vaccine appears to rely on
a reorientation of the immune response toward a Th1 type, with
the production of IFNγ and the activation of CD8+ T cells, and
the lack of CD8+ completely abrogate the therapeutic vaccine
4 Human Vaccines Volume 7 issue 11
effect.59,60 It is thus of key interest that the same antigens and vac-
cine formulations may be used for both the prevention of Chagas
disease as well as for the therapy of an ongoing infection, making
these a very flexible tool for disease control.
Given these promising results in mouse models, a few of
these vaccine candidates have been evaluated in dogs. These
are considered an excellent animal model, which presents a
disease progression much more similar to that of humans than
mice.61-63 In addition, as mentioned above, a veterinary vaccine
for the control of canine Chagas disease would be very helpful
for the control of this important parasite reservoir, as well as for
the care of pets which are increasingly found to be infected by
T. cruzi.16 Thus, immunization with a live attenuated T. cruzi
strain was found to significantly reduce natural T. cruzi infec-
tion in dogs in a large field study, as assessed by xenodiagnosis.64
Similarly, as observed in mice, immunization of dogs with T.
rangeli was able to reduce their infectiveness to triatomine vec-
tors following experimental infection, although all dogs devel-
oped parasitemia and it is unclear if they were protected from
further disease development.65 More recently, immunization
with four doses of TcVac1, based on a combination of DNA vac-
cines encoding TcG1, TcG2 and TcG4, dog Il-12 and GM-CSF
was evaluated.66 Immunization induced an increase in anti-
body levels, together with some increase in IFNγ and CD8+ T
cells, confirming the immunogenicity of the vaccine. However,
immunized dogs presented only a small decrease in infectiv-
ity to triatomines, as well as in cardiac arrhythmias., and the
inflammatory reaction in the heart appeared unchanged com-
pared with control dogs. Importantly, the density of necrosis,
inflammatory foci and amastigote nests (parasite burden) were
increased in vaccinated dogs, as well as their mortality, suggest-
ing a limited efficacy of this vaccine.66
As in mice, the therapeutic administration of two doses of DNA
vaccines encoding TSA-1 and Tc24 during the acute phase has
also been tested in dogs and EKG recording indicated a decrease
in the severity of cardiac arrythmias67 (Quijano-Hernandez et
al. unpublished data). Importantly, this DNA vaccine can also
reduce the density of amastigote nests, cardiac arrhythmias and
mortality when used as a preventive vaccine in dogs, emphasiz-
ing the potential of this formualtion (Quijano-Hernandez et al.
unpublished data). Taken together, these encouraging studies set
the basis for further evaluation of both preventive and therapeu-
tic vaccines in this animal model, which will be key to further
our understanding of T. cruzi control in non-murine models.
on antigens from the trans-sialidase family, but while such epit-
ope-specific CD8+ T cells are important for T. cruzi control, they
only provide strain-specific protection.50 Importantly, mice toler-
ized for these dominant epitopes are still able to control acute T.
cruzi infection through the activation of CD8+ T cells specific
for unknown alternate epitopes.51 Identifying these epitopes may
thus lead to additional vaccine antigens that could complement
trans-sialidase proteins.
Indeed, as noted before,19 a very limited variety of T. cruzi
antigens have been evaluated so far, and some studies have
focused on the identification of novel antigens. Thus for example,
the antigens TcG1, TcG2 and TcG4, corresponding to a puta-
tive ribosomal protein and hypothetical proteins, respectively,
have been recently identified and immunization with DNA
vaccines encoding all three antigens followed by recombinant
protein boosts (TcVac2) was found able to induce potent anti-
body, CD8+ T cell and cytokine responses and protected mice
Table 2. therapeutic vaccines against T. cruzi infection evaluated in mice
Formulation Antigens
Immune
response
Parasitemia
End points Heart
parasite burden
SurvivalHeart HistopathologyReference
Recombinant
protein
rSa85-1.1
increased
antibody levels
NRNR NRNo exacerbation 58
dNa
tSa-1, tc24,
and tc52
increased iFNγ
and cd8+
decreaseddecreasedincreased decreased inflammation
57, 59, 106,
60
aSp-2 like
clone 9 and tS
NRNo effect
No effectNo effect No effect106
NR: Not reported.

Page 5

©2011 Landes Bioscience.
Do not distribute.
be an attractive complement to vector control, to help mitigate the
rising problem of insecticide resistance in some vector species.
Second, a major concern with Chagas disease has been the
fear of exacerbating disease by stimulating autoimmunity
through vaccination instead of providing protection,20,21,70 which
has considerably slowed down vaccine research. Importantly, this
situation is very similar to that of several other pathogens for
which there is also concern about vaccine-increased susceptibil-
ity to infection.71,72 A typical example is dengue virus, in which
case antibody-dependent enhancement (ADE) caused by weakly
neutralizing but cross-reactive antibodies is believed to favor a
secondary infection by a heterologous serotype of the virus.
While the mechanisms of these processes are still not understood
in detail, these aspects are carefully taken into consideration in
vaccine development to avoid such adverse effects.73,74 In the case
of T. cruzi, there is now a large body of evidence indicating that
disease progression is due to parasite persistence, and autoim-
mune reactions are rather a consequence of cardiac tissue damage
than the main mechanism driving disease progression.75-78 Also,
several independent pre-clinical vaccine studies such as those
highlighted above have repeatedly shown that the induction of a
strong cellular immune response allows for a reduction in para-
site burden as well as in cardiac tissue inflammation, rather than
exacerbation of disease (Tables 1 and 2). Taken together, these
data clearly suggest that both preventive of therapeutic vaccina-
tion against T. cruzi are safe and do not lead to autoimmune
reactions. However, it is also clear that more extensive evaluations
www.landesbioscience.com Human Vaccines 5
are needed. Thus, severe adverse effects including autoimmunity
will need to be closely monitored in future vaccine trials, but do
not justify further delaying vaccine development.
Another major issue is the design and feasibility of clinical
trials. Indeed, chronic Chagas disease takes years to develop,
leading some authors to argue that trials would need to last at
least 20 y to see any significant effect.68 In addition, given the
low incidence of disease in most countries where intensive vec-
tor control has been implemented, randomized controlled trials
would need to include a very large number of volunteers to allow
for the detection of a modest vaccine efficacy. Again, these issues
are not specific to T. cruzi vaccines and have been raised for other
chronic diseases including TB or HIV-AIDS.79,80 It is clear that
they certainly complicate trial design, but it does not mean that
such trials are not feasible. A therapeutic vaccine trial may be
easier to design than a preventive vaccine trial, although as in the
case of HIV trials it would be necessary to administer the vaccine
together with conventional drug treatment, which would make
it more difficult to observed a moderate vaccine effect. In any
case, novel trial design such as adaptive trial designs may help to
reduce duration and number of volunteers needed, while ensur-
ing the scientific quality and integrity of such trials.81 A correlate
problem to trial design is that of the lack of widely accepted bio-
markers to evaluate vaccine efficacy. Thus, neither the end-points
to be used in a clinical trial, nor the expected target product
profile are easily defined. As illustrated above, commonly used
markers in animal studies, which include cardiac tissue dam-
age, parasite burden in the heart, disease reactivation following
immunosupression or survival, are too invasive and/or unethical
to be used in clinical trials. Also, there is some debate about the
need for a vaccine to be able to provide sterile immunity, lead-
ing some authors to discuss the usefulness of a vaccine unable
to reach this end-point. However, it seems that while desirable,
sterile immunity may not be a necessary target. Indeed, disease
progression to Chagasic congestive cardiomyopathy and cardiac
failure is what is causing the major morbidity of Chagas disease
and its economic cost. Thus, preventing or at least delaying the
onset of congestive cardiomyopathy may confer sufficient value
to a T. cruzi vaccine. Thus, clinical indicators such as EKG or
echocardiography may provide valuable end-points to evaluate
progressive cardiac damage and vaccine efficacy.82,83 Also, such
clinical indicators may be able to detect difference in disease pro-
gression rate within a reasonable time, as suggested by studies
in dogs66,67,84 and in patient follow-up evaluations.82,83,85 In that
respect, much can be learned from the current “BENEFIT” trial,
which is evaluating the efficacy of drug treatment in chronic-
stage patients. Nonetheless, ongoing efforts for the identification
of additional biomarkers are underway, including electrocar-
diographic, immunologic and biochemical markers, which may
greatly improve and facilitate the monitoring of Chagas disease
progression and thus vaccine evaluation.86-92
As indicated above, a successful vaccine will need to induce
a strong cellular immune response, with the activation of CD8+
cytotoxic T cells in order to effectively control T. cruzi parasites.
This requirement places some constraints on the formulations
and types of vaccines that may be used, since such a response
Perspectives and Challenges to T. cruzi Vaccine
Development
While many of the studies highlighted above provide proof-of-
concept data on the potential of a preventive or therapeutic vac-
cine to control a T. cruzi infection by at least decreasing parasite
burden, cardiac tissue inflammation and damage or increas-
ing survival, a number of important challenges and key ques-
tions remain to be answered for further vaccine development to
proceed.
First of all, some authors have argued against the rationale
presented above about the actual need for a vaccine.68,69 Indeed,
as transmission of the disease has been effectively controlled by
vector control interventions in large parts of Latin America, the
population at risk of infection has been dramatically reduced.
Thus, there seem to be no population left to vaccinate. However,
as we briefly mentioned, it is important to stress that domiciliated
Triatoma infestans and Rhodnius prolixus are the only vector species
which have been effectively controlled (although partially as insec-
ticide resistance has now been detected in T. infestans), and that
many autochthonous triatomine species are much more difficult to
control and will remain responsible for significant T. cruzi trans-
mission to humans, although at lower levels. Furthermore, the
cost-effectiveness of a vaccine has been demonstrated even when
there is a low prevalence of T. cruzi infection (about 1%10), which
is what can be expected to remain after such vector control efforts.
A vaccine would thus definitively be an asset for populations still at
risk, or for infected patients in the case of a therapeutic vaccine. In
addition, a veterinary vaccine targeting domestic reservoirs would

Page 6

©2011 Landes Bioscience.
Do not distribute.
effects of drugs used to treat Chagas’ disease (American
trypanosomiasis). Hum Exp Toxicol 2006; 25:471-
9; PMID:16937919; DOI:10.1191/0960327106
het653oa.
Jackson Y, Alirol E, Getaz L, Wolff H, Combescure
C, Chappuis F. Tolerance and safety of nifurti-
mox in patients with chronic chagas disease. Clin
Infect Dis 2010; 51:69-75; PMID:20932171;
DOI:10.1086/656917.
Muratore CA, Batista Sa LA, Chiale PA, Eloy R,
Tentori MC, Escudero J, et al. Implantable cardioverter
defibrillators and Chagas’ disease: results of the ICD
Registry Latin America. Europace 2009; 11:164-8;
PMID:19056745; DOI:10.1093/europace/eun325.
Castillo-Riquelme M, Guhl F, Turriago B, Pinto N,
Rosas F, Martinez MF, et al. The costs of preventing and
treating chagas disease in Colombia. PLoS Negl Trop
Dis 2008; 2:336; PMID:19015725; DOI:10.1371/
journal.pntd.0000336.
Dias JC, Silveira AC, Schofield CJ. The impact of
Chagas disease control in Latin America: a review. Mem
Inst Oswaldo Cruz 2002; 97:603-12; PMID:12219120;
DOI:10.1590/S0074-02762002000500002.
Schofield CJ, Jannin J, Salvatella R. The future of
Chagas disease control. Trends Parasitol 2006; 22:583-
8; PMID:17049308; DOI:10.1016/j.pt.2006.09.011.
10. Lee BY, Bacon KM, Connor DL, Willig AM, Bailey
RR. The potential economic value of a Trypanosoma
cruzi (Chagas disease) vaccine in Latin America.
PLoS Negl Trop Dis 2010; 4:916; PMID:21179503;
DOI:10.1371/journal.pntd.0000916.
11. Montenegro VM, Jimenez M, Pinto Dias JC,
Zeledon R. Chagas disease in dogs from endemic
areas of Costa Rica. Mem Inst Oswaldo Cruz 2002;
97:491-4; PMID:12118277; DOI:10.1590/S0074-
02762002000400006.
90; PMID:21243314;
879X2011007500005.
6 Human Vaccines Volume 7 issue 11
In conclusion, after years of neglect, it appears that vaccine
development against T. cruzi is finally coming of age. Abundant
mice studies provide strong proof-of-concept data on the feasi-
bility of controlling T. cruzi infection with both preventive and
therapeutic vaccines, and it is clear that such vaccines would be
extremely helpful for Chagas disease control. Importantly, the
emergence of novel international partnerships bringing together
academics and manufacturers is expected to provide a renewed
momentum to vaccine development against neglected diseases.93
While many challenges remain to be addressed, including tech-
nical, ethical and logistical issues, advances in our understand-
ing of Chagas disease pathogenesis and in vaccine development
against other diseases are providing some clues about how to
address these. Encouraging results from dog trials suggest that
a veterinary vaccine could be developed in the short-term, which
would provide further support for the development of a human
vaccine.
is harder to induce than a humoral response. In addition, for a
vaccine to be efficiently distributed in remote rural areas with
limited health infrastructure, a very stable formulation would be
a key advantage.
A therapeutic vaccine targeting infected adult patients in
either the indeterminate or symptomatic chronic phase could
be administered through health centers, alone or in combina-
tion with conventional drug therapy, to prevent or delay disease
progression. Such a vaccine would be of interest in most Latin
American countries where the disease is endemic, as well as in
non-endemic countries with large populations of migrants from
this region. Alternatively, a preventive vaccine could be aimed at
both children and adult populations living in endemic areas, even
where vectorial transmission has been considerably reduced, to
further reduce the incidence of new cases of Chagasic cardiomy-
opathy. Such a vaccine could eventually be integrated into estab-
lished vaccination programs.
References
1. Rassi A Jr, Rassi A, Marin-Neto JA. Chagas dis-
ease. Lancet 2010; 375:1388-402; PMID:20399979;
DOI:10.1016/S0140-6736(10)60061-X.
Pérez-Molina JA, Perez-Ayala A, Moreno S, Fernandez-
Gonzalez MC, Zamora J, Lopez-Velez R. Use of benz-
nidazole to treat chronic Chagas’ disease: a systematic
review with a meta-analysis. J Antimicrob Chemother
2009; 64:1139-47; PMID:19819909; DOI:10.1093/
jac/dkp357.
Lescure FX, Le Loup G, Freilij H, Develoux M,
Paris L, Brutus L, et al. Chagas disease: changes in
knowledge and management. Lancet Infect Dis 2010;
10:556-70; PMID:20670903; DOI:10.1016/S1473-
3099(10)70098-0.
Castro JA, de Mecca MM, Bartel LC. Toxic side
2.
3.
4.
5.
6.
7.
8.
9.
12. Crisante G, Rojas A, Teixeira MM, Anez N. Infected
dogs as a risk factor in the transmission of human
Trypanosoma cruzi infection in western Venezuela.
Acta Trop 2006; 98:247-54; PMID:16797466;
DOI:10.1016/j.actatropica.2006.05.006.
13. Estrada-Franco JG, Bhatia V, Diaz-Albiter H, Ochoa-
Garcia L, Barbabosa A, Vazquez-Chagoyan JC, et al.
Human Trypanosoma cruzi infection and seropositivity
in dogs, Mexico. Emerg Infect Dis 2006; 12:624-30;
PMID:16704811.
14. Gürtler RE, Cecere MC, Lauricella MA, Cardinal
MV, Kitron U, Cohen JE. Domestic dogs and
cats as sources of Trypanosoma cruzi infection in
rural northwestern Argentina. Parasitology 2007;
134:69-82; PMID:17032467;
S0031182006001259.
15. Reithinger R, Ceballos L, Stariolo R, Davies CR,
Gurtler RE. Extinction of experimental Triatoma infes-
tans populations following continuous exposure to dogs
wearing deltamethrin-treated collars. Am J Trop Med
Hyg 2006; 74:766-71; PMID:16687678.
16. Kjos SA, Snowden KF, Craig TM, Lewis B, Ronald
N, Olson JK. Distribution and characterization of
canine Chagas disease in Texas. Vet Parasitol 2008;
152:249-56; PMID:18255233; DOI:10.1016/j.vet-
par.2007.12.021.
17. ,Garg N, Bhatia V. Current status and future prospects
for a vaccine against American trypanosomiasis. Expert
Rev Vaccines 2005; 4:867-80; PMID:16372882;
DOI:10.1586/14760584.4.6.867.
18. Cazorla SI, Frank FM, Malchiodi EL. Vaccination
approaches against Trypanosoma cruzi infection. Expert
Rev Vaccines 2009; 8:921-35; PMID:19538117;
DOI:10.1586/erv.09.45.
19. Dumonteil E. Vaccine development against Trypanosoma
cruzi and Leishmania species in the post-genomic era.
Infect Genet Evol 2009; In press; PMID:19805015;
DOI:10.1016/j.meegid.2009.02.009.
20. Kierszenbaum F. Chagas’ disease and the autoimmu-
nity hypothesis. Clin Microbiol Rev 1999; 12:210-23;
PMID:10194457.
21. Tarleton RL, Zhang L. Chagas Disease Etiology:
Autoimmunity or Parasite Persistence? Parasitol Today
1999; 15:94-9; PMID:10322321; DOI:10.1016/
S0169-4758(99)01398-8.
22. Zhang L, Tarleton RL. Parasite persistence corre-
lates with disease severity and localization in chron-
ic Chagas’ disease. J Infect Dis 1999; 180:480-6;
PMID:10395865; DOI:10.1086/314889.
23. DosReis GA. Evasion of immune respons-
es by Trypanosoma cruzi, the etiological agent of
Chagas disease. Braz J Med Biol Res 2011; 44:84-
DOI:10.1017/
DOI:10.1590/S0100-
24. Ouaissi A, Da Silva AC, Guevara AG, Borges M,
Guilvard E. Trypanosoma cruzi-Induced Host Immune
System Dysfunction: A Rationale for Parasite
Immunosuppressive Factor(s) Encoding Gene Targeting.
J Biomed Biotechnol 2001; 1:11-7; PMID:12488621;
DOI:10.1155/S1110724301000055.
25. Alba Soto CD, Solana ME, Poncini CV, Pino-Martinez
AM, Tekiel V, Gonzalez-Cappa SM. Dendritic cells
devoid of IL-10 induce protective immunity against
the protozoan parasite Trypanosoma cruzi. Vaccine
2010; 28:7407-13; PMID:20850535; DOI:10.1016/j.
vaccine.2010.08.105.
26. Ouaissi A, Guilvard E, Delneste Y, Caron G, Magistrelli
G, Herbault N, et al. The Trypanosoma cruzi Tc52-
released protein induces human dendritic cell matu-
ration, signals via Toll-like receptor 2, and confers
protection against lethal infection. J Immunol 2002;
168:6366-74; PMID:12055254.
27. Alvarez MN, Peluffo G, Piacenza L, Radi R.
Intraphagosomal peroxynitrite as a macrophage-
derived cytotoxin against internalized Trypanosoma
cruzi: consequences for oxidative killing and role of
microbial peroxiredoxins in infectivity. J Biol Chem
2011; 286:6627-40; PMID:21098483; DOI:10.1074/
jbc.M110.167247.
28. Padilla AM, Bustamante JM, Tarleton RL. CD8+ T cells
in Trypanosoma cruzi infection. Curr Opin Immunol
2009; 21:385-90; PMID:19646853; DOI:10.1016/j.
coi.2009.07.006.
29. Parodi C, Padilla AM, Basombrio MA. Protective
immunity against Trypanosoma cruzi. Mem Inst
Oswaldo Cruz 2009; 104:288-94; PMID:19753487;
DOI:10.1590/S0074-02762009000900038.
30. de Alencar BC, Persechini PM, Haolla FA, de Oliveira
G, Silverio JC, Lannes-Vieira J, et al. Perforin and
gamma interferon expression are required for CD4+ and
CD8+ T-cell-dependent protective immunity against a
human parasite, Trypanosoma cruzi, elicited by heter-
ologous plasmid DNA prime-recombinant adenovirus
5 boost vaccination. Infect Immun 2009; 77:4383-95;
PMID:19651871; DOI:10.1128/IAI.01459-08.
31. Sardinha LR, Mosca T, Elias RM, do Nascimento RS,
Goncalves LA, Bucci DZ, et al. The liver plays a major
role in clearance and destruction of blood trypomasti-
gotes in Trypanosoma cruzi chronically infected mice.
PLoS Negl Trop Dis 2010; 4:578; PMID:20052269;
DOI:10.1371/journal.pntd.0000578.
32. Collins MH, Craft JM, Bustamante JM, Tarleton RL.
Oral exposure to Trypanosoma cruzi elicits a systemic
CD8+ T cell response and protection against hetero-
topic challenge. Infect Immun 2011; 79:3397-406;
PMID:21628516; DOI:10.1128/IAI.01080-10.

Page 7

©2011 Landes Bioscience.
Do not distribute.
4:841; PMID:20957201; DOI:10.1371/journal.
pntd.0000841.
56. Ouaissi A, Aguirre T, Plumas Marty B, Piras M,
Schoneck R, Gras Masse H, et al. Cloning and
sequencing of a 24-kDa Trypanosoma cruzi specific
antigen released in association with membrane vesicles
and defined by a monoclonal antibody. Biol Cell
1992; 75:11-7; PMID:1381253; DOI:10.1016/0248-
4900(92)90119-L.
57. Dumonteil
E, Escobedo-Ortegon
Rodriguez N, Ramirez-Sierra MJ, Arjona-Torres A.
Immunotherapy of Trypanosoma cruzi infection with
DNA vaccines in mice. Infect Immun 2004; 72:46-
53; PMID:14688079; DOI:10.1128/IAI.72.1.46-
53.2004.
58. Duthie MS, Kahn M, Zakayan A, White M, Kahn SJ.
Parasite-induced chronic inflammation is not exacer-
bated by immunotherapy before or during Trypanosoma
cruzi Infection. Clin Vaccine Immunol 2007; 14:1005-
12; PMID:17538117; DOI:10.1128/CVI.00087-07.
59. Zapata-Estrella H, Hummel-Newell C, Sanchez-
Burgos G, Escobedo-Ortegon J, Ramirez-Sierra MJ,
Arjona-Torres A, et al. Control of Trypanosoma cruzi
infection and changes in T cell populations induced
by a therapeutic DNA vaccine in mice. Immunol Lett
2006; 103:186-91; PMID:16378645; DOI:10.1016/j.
imlet.2005.11.015.
60. Limon-Flores AY, Cervera-Cetina R, Tzec-Arjona JL,
Ek-Macias L, Sanchez-Burgos G, Ramirez-Sierra MJ,
et al. Effect of a combination DNA vaccine for the
prevention and therapy of Trypanosoma cruzi infec-
tion in mice: role of CD4+ and CD8+ T cells. Vaccine
2010; 28:7414-9; PMID:20850536; DOI:10.1016/j.
vaccine.2010.08.104.
vector vaccination can successfully control Trypanosoma
cruzi infection. Infect Immun 2005; 73:7356-65;
PMID:16239534; DOI:10.1128/IAI.73.11.7356-
65.2005.
www.landesbioscience.com Human Vaccines 7
61. Barr SC, Gossett KA, Klei TR. Clinical, clinicopatho-
logic and parasitologic observations of trypanosomiasis
in dogs infected with North American Trypanosoma
cruzi isolates. Am J Vet Res 1991; 52:954-60;
PMID:1909105.
62. Guedes PM, Veloso VM, Tafuri WL, Galvao LM,
Carneiro CM, Lana M, et al. The dog as model for
chemotherapy of the Chagas’ disease. Acta Trop 2002;
84:9-17; PMID:12387906; DOI:10.1016/S0001-
706X(02)00139-0.
63. Cruz-Chan
JV, Bolio-González
Flores R, Ramirez-Sierra MJ, Quijano-Hernandez
IA, Dumonteil E. Immunopathology of natural
Trypanosoma cruzi infection in dogs. Vet Parasitol
2009; 162:151-5; PMID:19324497; DOI:10.1016/j.
vetpar.2009.02.024.
64. Basombrio MA, Segura MA, Mora MC, Gomez L.
Field trial of vaccination against American trypanoso-
miasis (Chagas’ disease) in dogs. Am J Trop Med Hyg
1993; 49:143-51; PMID:8352387.
65. Basso B, Castro I, Introini V, Gil P, Truyens C,
Moretti E. Vaccination with Trypanosoma rangeli
reduces the infectiousness of dogs experimental-
ly infected with Trypanosoma cruzi. Vaccine 2007;
25:3855-8; PMID:17349724; DOI:10.1016/j.vac-
cine.2007.01.114.
66. Aparicio-Burgos JE, Ochoa-Garcia L, Zepeda-
Escobar JA, Gupta S, Dhiman M, Martinez JS, et al.
Testing the Efficacy of a Multi-Component DNA-
Prime/DNA-Boost Vaccine against Trypanosoma
cruzi Infection in Dogs. PLoS Negl Trop Dis 2011;
5:1050; PMID:21625470; DOI:10.1371/journal.
pntd.0001050.
67. Quijano-Hernandez
IA,
Rodríguez-Buenfil JC, Ramirez-Sierra MJ, Dumonteil
E. Therapeutic DNA vaccine against Trypansomoma
cruzi in dogs: a pilot clinical trial. Ann NY Acad Sci
2008; 1149:343-6; PMID:19120245; DOI:10.1196/
annals.1428.098.
68. Camargo EP. Perspectives of vaccination in Chagas
disease revisited. Mem Inst Oswaldo Cruz 2009;
104:275-80; PMID:19753485; DOI:10.1590/S0074-
02762009000900036.
69. Junqueira C, Caetano B, Bartholomeu DC, Melo
MB, Ropert C, Rodrigues MM, et al. The endless race
between Trypanosoma cruzi and host immunity: les-
sons for and beyond Chagas disease. Expert Rev Mol
Med 2010; 12:29; PMID:20840799; DOI:10.1017/
S1462399410001560.
70. Engman DM, Leon JS. Pathogenesis of Chagas
heart disease: role of autoimmunity. Acta Trop 2002;
81:123-32; PMID:11801219; DOI:10.1016/S0001-
706X(01)00202-9.
71. Thomas S, Redfern JB, Lidbury BA, Mahalingam
S. Antibody-dependent enhancement and vaccine
development. Expert Rev Vaccines 2006; 5:409-12;
PMID:16989620; DOI:10.1586/14760584.5.4.409.
72. Huisman W, Martina BE, Rimmelzwaan GF,
Gruters RA, Osterhaus AD. Vaccine-induced
enhancement of viral infections. Vaccine 2009;
27:505-12; PMID:19022319; DOI:10.1016/j.vac-
cine.2008.10.087.
73. Guy B, Chanthavanich P, Gimenez S, Sirivichayakul
C, Sabchareon A, Begue S, et al. Evaluation by
flow cytometry of antibody-dependent enhancement
(ADE) of dengue infection by sera from Thai children
immunized with a live-attenuated tetravalent dengue
vaccine. Vaccine 2004; 22:3563-74; PMID:15315835;
DOI:10.1016/j.vaccine.2004.03.042.
74. Dejnirattisai W, Jumnainsong A, Onsirisakul N, Fitton
P, Vasanawathana S, Limpitikul W, et al. Cross-
reacting antibodies enhance dengue virus infection in
humans. Science 2010; 328:745-8; PMID:20448183;
DOI:10.1126/science.1185181.
75. Tarleton RL. Parasite persistence in the aetiol-
ogy of Chagas disease. Int J Parasitol 2001;
31:550-4; PMID:11334941; DOI:10.1016/S0020-
7519(01)00158-8.
ME, Colin-
Bolio-González ME,
48. Martin DL, Weatherly DB, Laucella SA, Cabinian MA,
Crim MT, Sullivan S, et al. CD8+ T-Cell responses
to Trypanosoma cruzi are highly focused on strain-
variant trans-sialidase epitopes. PLoS Pathog 2006;
2:77; PMID:16879036;
ppat.0020077.
49. Tzelepis F, de Alencar BC, Penido ML, Claser C,
Machado AV, Bruna-Romero O, et al. Infection with
Trypanosoma cruzi restricts the repertoire of parasite-
specific CD8+ T cells leading to immunodominance. J
Immunol 2008; 180:1737-48; PMID:18209071.
50. Haolla FA, Claser C, de Alencar BC, Tzelepis F, de
Vasconcelos JR, de Oliveira G, et al. Strain-specific
protective immunity following vaccination against
experimental Trypanosoma cruzi infection. Vaccine
2009; 27:5644-53; PMID:19635607; DOI:10.1016/j.
vaccine.2009.07.013.
51. Rosenberg CS, Martin DL, Tarleton RL. CD8+ T cells
specific for immunodominant trans-sialidase epitopes
contribute to control of Trypanosoma cruzi infection
but are not required for resistance. J Immunol 2010;
185:560-8; PMID:20530265; DOI:10.4049/jimmu-
nol.1000432.
52. Bhatia V, Garg NJ. Previously unrecognized vaccine
candidates control Trypanosoma cruzi infection and
immunopathology in mice. Clin Vaccine Immunol
2008; 15:1158-64; PMID:18550728; DOI:10.1128/
CVI.00144-08.
53. Gupta S, Garg NJ. Prophylactic efficacy of TcVac2
against Trypanosoma cruzi in mice. PLoS Negl Trop Dis
2010; 4:797; PMID:20706586; DOI:10.1371/journal.
pntd.0000797.
54. Tekiel V, Alba-Soto CD, Gonzalez Cappa SM, Postan
M, Sanchez DO. Identification of novel vaccine candi-
dates for Chagas’ disease by immunization with sequen-
tial fractions of a trypomastigote cDNA expression
library. Vaccine 2009; 27:1323-32; PMID:19162108;
DOI:10.1016/j.vaccine.2008.12.056.
55. García EA, Ziliani M, Aguero F, Bernabo G, Sanchez
DO, Tekiel V. TcTASV: a novel protein family in
Trypanosoma cruzi identified from a subtractive trypo-
mastigote cDNA library. PLoS Negl Trop Dis 2010;
DOI:10.1371/journal.
J, Reyes-
33. Basombrio MA. Trypanosoma cruzi: partial prevention of
the natural infection of guinea pigs with a killed parasite
vaccine. Exp Parasitol 1990; 71:1-8; PMID:2113004;
DOI:10.1016/0014-4894(90)90002-T.
34. Basombrío MA, Besuschio S. Trypanosoma cruzi culture
used as vaccine to prevent chronic Chagas’ disease in
mice. Infect Immun 1982; 36:351-6; PMID:6804390.
35. Zuñiga C, Palau T, Penin P, Gamallo C, de Diego JA.
Protective effect of Trypanosoma rangeli against infec-
tions with a highly virulent strain of Trypanosoma cruzi.
Trop Med Int Health 1997; 2:482-7; PMID:9217704;
DOI:10.1111/j.1365-3156.1997.tb00171.x.
36. Basso B, Cervetta L, Moretti E, Carlier Y, Truyens C.
Acute Trypanosoma cruzi infection: IL-12, IL-18, TNF,
sTNFR and NO in T. rangeli-vaccinated mice. Vaccine
2004; 22:1868-72; PMID:15121297; DOI:10.1016/j.
vaccine.2003.11.013.
37. Basso B, Moretti E, Fretes R. Vaccination with epimasti-
gotes of different strains of Trypanosoma rangeli protects
mice against Trypanosoma cruzi infection. Mem Inst
Oswaldo Cruz 2008; 103:370-4; PMID:18660992;
DOI:10.1590/S0074-02762008000400010.
38. Zuñiga E, Motran C, Montes CL, Diaz FL, Bocco JL,
Gruppi A. Trypanosoma cruzi-induced immunosup-
pression: B cells undergo spontaneous apoptosis and
lipopolysaccharide (LPS) arrests their proliferation
during acute infection. Clin Exp Immunol 2000;
119:507-15; PMID:10691924; DOI:10.1046/j.1365-
2249.2000.01150.x.
39. Taibi A, Plumas-Marty B, Guevara-Espinoza A,
Schöneck R, Pessoa H, Loyens M, et al. Trypanosoma
cruzi: immunity-induced in mice and rats by trypomas-
tigote excretory-secretory antigens and identification
of a peptide sequence containing a T cell epitope with
protective activity. J Immunol 1993; 151:2676-89;
PMID:7689612.
40. Wrightsman RA, Miller MJ, Saborio JL, Manning JE.
Pure paraflagellar rod protein protects mice against
Trypanosoma cruzi infection. Infect Immun 1995;
63:122-5; PMID:7806347.
41. Miller MJ, Wrightsman RA, Manning JE. Trypanosoma
cruzi: protective immunity in mice immunized with
paraflagellar rod proteins is associated with a T-helper
type 1 response. Exp Parasitol 1996; 84:156-67;
PMID:8932765; DOI:10.1006/expr.1996.0101.
42. Frank FM, Petray PB, Cazorla SI, Munoz MC, Corral
RS, Malchiodi EL. Use of a purified Trypanosoma cruzi
antigen and CpG oligodeoxynucleotides for immuno-
protection against a lethal challenge with trypomas-
tigotes. Vaccine 2003; 22:77-86; PMID:14604574;
DOI:10.1016/S0264-410X(03)00541-3.
43. Crampton A, Vanniasinkam T. Parasite vac-
cines: the new generation. Infect Genet Evol 2007;
7:664-73; PMID:17702669;
meegid.2007.06.004.
44. Dumonteil E. DNA vaccines against protozoan
parasites: opportunities and challenges. J Biomed
Biotechnol 2007; 2007:90520; PMID:17710244;
DOI:10.1155/2007/90520.
45. Nogueira RT, Nogueira AR, Pereira MC, Rodrigues
MM, Galler R, Bonaldo MC. Biological and immu-
nological characterization of recombinant Yellow Fever
17D viruses expressing a Trypanosoma cruzi Amastigote
Surface Protein-2 CD8+ T cell epitope at two dis-
tinct regions of the genome. Virol J 2011; 8:127;
PMID:21418577; DOI:10.1186/1743-422X-8-127.
46. Takayama E, Ono T, Carnero E, Umemoto S,
Yamaguchi Y, Kanayama A, et al. Quantitative and qual-
itative features of heterologous virus-vector-induced
antigen-specific CD8+ T cells against Trypanosoma
cruzi infection. Int J Parasitol 2010; 40:1549-61;
PMID:20620143; DOI:10.1016/j.ijpara.2010.05.011.
47. Miyahira Y, Takashima Y, Kobayashi S, Matsumoto Y,
Takeuchi T, Ohyanagi-Hara M, et al. Immune respons-
es against a single CD8+-T-cell epitope induced by virus
DOI:10.1016/j.

Page 8

©2011 Landes Bioscience.
Do not distribute.
infection: importance of a TLR9 agonist and CD4+
T cells. Vaccine 2007; 25:6018-27; PMID:17629597;
DOI:10.1016/j.vaccine.2007.05.022.
96. Hoft DF, Eickhoff CS, Giddings OK, Vasconcelos
JR, Rodrigues MM. Trans-sialidase recombinant pro-
tein mixed with CpG motif-containing oligodeoxy-
nucleotide induces protective mucosal and systemic
Trypanosoma cruzi immunity involving CD8+ CTL
and B cell-mediated cross-priming. J Immunol 2007;
179:6889-900; PMID:17982080.
97. Giddings OK, Eickhoff CS, Sullivan NL, Hoft DF.
Intranasal vaccinations with the trans-sialidase antigen
plus CpG Adjuvant induce mucosal immunity protec-
tive against conjunctival Trypanosoma cruzi challenges.
Infect Immun 2010; 78:1333-8; PMID:20048046;
DOI:10.1128/IAI.00278-09.
2010; 4:826; PMID:20877635; DOI:10.1371/journal.
pntd.0000826.
88. Lima-Costa MF, Cesar CC, Peixoto SV, Ribeiro AL.
Plasma B-type natriuretic peptide as a predictor of mor-
tality in community-dwelling older adults with Chagas
disease: 10-year follow-up of the Bambui Cohort
Study of Aging. Am J Epidemiol 2010; 172:190-6;
PMID:20581155; DOI:10.1093/aje/kwq106.
8 Human Vaccines Volume 7 issue 11
98. Cazorla SI, Frank FM, Becker PD, Arnaiz M, Mirkin
GA, Corral RS, et al. Redirection of the immune
response to the functional catalytic domain of the cys-
tein proteinase cruzipain improves protective immunity
against Trypanosoma cruzi infection. J Infect Dis 2010;
202:136-44; PMID:20497050; DOI:10.1086/652872.
99. Eickhoff CS, Giddings OK, Yoshida N, Hoft DF.
Immune responses to gp82 provide protection against
mucosal Trypanosoma cruzi infection. Mem Inst
Oswaldo Cruz 2010; 105:687-91; PMID:20835618;
DOI:10.1590/S0074-02762010000500015.
100. Machado AV, Cardoso JE, Claser C, Rodrigues MM,
Gazzinelli RT, Bruna-Romero O. Long-term protec-
tive immunity induced against Trypanosoma cruzi
infection after vaccination with recombinant adeno-
viruses encoding amastigote surface protein-2 and
trans-sialidase. Hum Gene Ther 2006; 17:898-908;
PMID:16972758; DOI:10.1089/hum.2006.17.898.
101. Duan X, Yonemitsu Y, Chou B, Yoshida K, Tanaka
S, Hasegawa M, et al. Efficient protective immu-
nity against Trypanosoma cruzi infection after nasal
vaccination with recombinant Sendai virus vector
expressing amastigote surface protein-2. Vaccine 2009;
27:6154-9; PMID:19712768; DOI:10.1016/j.vac-
cine.2009.08.026.
102. Garg N, Tarleton RL. Genetic immunization elic-
its antigen-specific protective immune responses and
decreases disease severity in Trypanosoma cruzi infection.
Infect Immun 2002; 70:5547-55; PMID:12228281;
DOI:10.1128/IAI.70.10.5547-55.2002.
103. Chou B, Hiromatsu K, Hisaeda H, Duan X, Imai
T, Murata S, et al. Genetic immunization based
on the ubiquitin-fusion degradation pathway against
Trypanosoma cruzi. Biochem Biophys Res Commun
2010; 392:277-82; PMID:20059980; DOI:10.1016/j.
bbrc.2009.12.166.
104. Eickhoff CS, Vasconcelos JR, Sullivan NL,
Blazevic A, Bruna-Romero O, Rodrigues MM, et
al. Co-administration of a plasmid DNA encoding
IL-15 improves long-term protection of a genetic vac-
cine against Trypanosoma cruzi. PLoS Negl Trop Dis
2011; 5:983; PMID:21408124; DOI:10.1371/journal.
pntd.0000983.
105. Rigato PO, de Alencar BC, de Vasconcelos JR,
Dominguez MR, Araujo AF, Machado AV, et al.
Heterologous plasmid DNA prime-recombinant
human adenovirus 5 boost vaccination generates a
stable pool of protective long-lived CD8(+) T effec-
tor memory cells specific for a human parasite,
Trypanosoma cruzi. Infect Immun 2011; 79:2120-30;
PMID:21357719; DOI:10.1128/IAI.01190-10.
106. Sanchez-Burgos G, Mezquita-Vega G, Escobedo-
Ortegon J, Ramirez-Sierra MJ, Arjona-Torres A,
Rodrigues MM, et al. Comparative efficacy of DNA
vaccines encoding various Trypanosoma cruzi antigens.
FEMS Microbiol Med Immunol 2007; 50:333-41;
DOI:10.1111/j.1574-695X.2007.00251.x.
89. Lorena VM, Lorena IM, Braz SC, Melo AS, Melo
MF, Melo MG, et al. Cytokine levels in serious car-
diopathy of Chagas disease after in vitro stimulation
with recombinant antigens from Trypanosoma cruzi.
Scand J Immunol 2010; 72:529-39; PMID:21044127;
DOI:10.1111/j.1365-3083.2010.02462.x.
90. Ndao M, Spithill TW, Caffrey R, Li H, Podust VN,
Perichon R, et al. Identification of novel diagnos-
tic serum biomarkers for Chagas’ disease in asymp-
tomatic subjects by mass spectrometric profiling. J
Clin Microbiol 2010; 48:1139-49; PMID:20071547;
DOI:10.1128/JCM.02207-09.
91. Wang Y, Moreira Mda C, Heringer-Walther S,
Ebermann L, Schultheiss HP, Wessel N, et al. Plasma
ACE2 activity is an independent prognostic marker in
Chagas’ disease and equally potent as BNP. J Card Fail
2010; 16:157-63; PMID:20142028; DOI:10.1016/j.
cardfail.2009.09.005.
92. Fabbro DL, Olivera V, Bizai ML, Denner S, Diez C,
Mancipar I, et al. Humoral immune response against
P2beta from Trypanosoma cruzi in persons with chronic
Chagas disease: its relationship with treatment against
parasites and myocardial damage. Am J Trop Med Hyg
2011; 84:575-80; PMID:21460013; DOI:10.4269/
ajtmh.2011; 10-0261.
93. Bottazzi ME, Dumonteil E, Valenzuela JG, Betancourt-
Cravioto M, Tapia-Conyer R, Hotez PJ. Bridging
the innovation gap for neglected tropical diseases in
Mexico: capacity building for the development of a
new generation of antipoverty vaccines. Bol Med Hosp
Infant Mex 2011; 68:130-8.
94. Araújo AF, de Alencar BC, Vasconcelos JR, Hiyane
MI, Marinho CR, Penido ML, et al. CD8+-T-cell-
dependent control of Trypanosoma cruzi infection in
a highly susceptible mouse strain after immuniza-
tion with recombinant proteins based on amastigote
surface protein 2. Infect Immun 2005; 73:6017-
25; PMID:16113322; DOI:10.1128/IAI.73.9.6017-
25.2005.
95. de Alencar BC, Araujo AF, Penido ML, Gazzinelli
RT, Rodrigues MM. Cross-priming of long lived
protective CD8+ T cells against Trypanosoma cruzi
76. Rocha MO, Ribeiro AL, Teixeira MM. Clinical manage-
ment of chronic chagas cardiomyopathy. Front Biosci
2003; 8:44-54; PMID:12456332; DOI:10.2741/926.
77. Hyland KV, Leon JS, Daniels MD, Giafis N, Woods
LM, Bahk TJ, et al. Modulation of autoimmunity
by treatment of an infectious disease. Infect Immun
2007; 75:3641-50; PMID:17485457; DOI:10.1128/
IAI.00423-07.
78. Marin-Neto JA, Cunha-Neto E, Maciel BC, Simoes
MV. Pathogenesis of chronic Chagas heart disease.
Circulation 2007; 115:1109-23; PMID:17339569;
DOI:10.1161/CIRCULATIONAHA.106.624296.
79. McKinnon LR, Card CM. HIV vaccine efficacy trials:
A brief history, and options for going forward. AIDS
Rev 2010; 12:209-17; PMID:21179185.
80. Padian NS, McCoy SI, Balkus JE, Wasserheit JN.
Weighing the gold in the gold standard: chal-
lenges in HIV prevention research. AIDS 2010;
24:621-35; PMID:20179575;
QAD.0b013e328337798a.
81. Bowalekar S. Adaptive designs in clinical trials.
Perspect Clin Res 2011; 2:23-7; PMID:21584178;
DOI:10.4103/2229-3485.76286.
82. Ribeiro AL, Rocha MO, Terranova P, Cesarano M,
Nunes MD, Lombardi F. T-wave amplitude variability
and the risk of death in Chagas Disease. J Cardiovasc
Electrophysiol 2011; In press; PMID:21235679;
DOI:10.1111/j.1540-8167.2010.02000.x.
83. Sarabanda AV, Marin-Neto JA. Predictors of mortality
in patients with Chagas’ cardiomyopathy and ventricu-
lar tachycardia not treated with implantable cardio-
verter-defibrillators. Pacing Clin Electrophysiol 2011;
34:54-62; PMID:20946310; DOI:10.1111/j.1540-
8159.2010.02896.x.
84. Barr SC, Warner KL, Kornreic BG, Piscitelli J, Wolfe
A, Benet L, et al. A cysteine protease inhibitor pro-
tects dogs from cardiac damage during infection by
Trypanosoma cruzi. Antimicrob Agents Chemother
2005; 49:5160-1; PMID:16304193; DOI:10.1128/
AAC.49.12.5160-1.2005.
85. Fernandes CD, Tiecher FM, Balbinot MM, Liarte
DB, Scholl D, Steindel M, et al. Efficacy of benzni-
dazol treatment for asymptomatic chagasic patients
from state of Rio Grande do Sul evaluated during a
three years follow-up. Mem Inst Oswaldo Cruz 2009;
104:27-32; PMID:19274372; DOI:10.1590/S0074-
02762009000100004.
86. Laucella SA, Mazliah DP, Bertocchi G, Alvarez MG,
Cooley G, Viotti R, et al. Changes in Trypanosoma cru-
zi-specific immune responses after treatment: surrogate
markers of treatment efficacy. Clin Infect Dis 2009;
49:1675-84; PMID:19877967; DOI:10.1086/648072.
87. Garcia-Alvarez A, Sitges M, Pinazo MJ, Regueiro-
Cueva A, Posada E, Poyatos S, et al. Chagas car-
diomyopathy: the potential of diastolic dysfunction
and brain natriuretic peptide in the early identi-
fication of cardiac damage. PLoS Negl Trop Dis
DOI:10.1097/