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Medicine » Infectious Diseases » "Leishmaniasis - Trends in Epidemiology, Diagnosis and Treatment", book edited by David M. Claborn, ISBN 978-953-51-1232-7, Published: March 19, 2014 under CC BY 3.0 license. © The Author(s).

Chapter 21

Novel Therapeutic Approaches to Leishmania Infection

By Levi H.C. Makala and Babak Baban
DOI: 10.5772/58167

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A glimpse of various antigens that have been used as vaccine candidates in past ten years (Adapted and modified with permission from Singh, and Sundar, 2012).
Figure 1. A glimpse of various antigens that have been used as vaccine candidates in past ten years (Adapted and modified with permission from Singh, and Sundar, 2012).
A conceptual model of IDO-mediated activation and effector T cell suppression following L. major infection. The model depicts interactions between IDO+ DCs, Tregs and naïve T cells that drive suppressive and non-suppressive outcomes under IDO-sufficient (+) and IDO-deficient (-) conditions in response to L. major infection. Induced IDO activity in DCs triggers cell stress responses blocks IL6 production by pDCs themselves, and by other cells (e.g. macrophages) capable of producing IL6. Under conditions of IDO ablation the same stimuli do not create suppression, and instead DCs stimulate naïve T cells, and express IL6, which converts Tregs to TH17 T cells or promotes TH17 differentiation from naïve CD4+ T cells (Adapted and modified from Makala, 2012).
Figure 2. A conceptual model of IDO-mediated activation and effector T cell suppression following L. major infection. The model depicts interactions between IDO+ DCs, Tregs and naïve T cells that drive suppressive and non-suppressive outcomes under IDO-sufficient (+) and IDO-deficient (-) conditions in response to L. major infection. Induced IDO activity in DCs triggers cell stress responses blocks IL6 production by pDCs themselves, and by other cells (e.g. macrophages) capable of producing IL6. Under conditions of IDO ablation the same stimuli do not create suppression, and instead DCs stimulate naïve T cells, and express IL6, which converts Tregs to TH17 T cells or promotes TH17 differentiation from naïve CD4+ T cells (Adapted and modified from Makala, 2012).
An illustration of the crystal structure for Indoleamine 2,3-dioxygenase (IDO) and chemical structures of its Inhibitors. A. bound to an inhibitor molecule.Indoleamine-2,3-dioxygenase inhibitors. Adapted from B. INCB024360 adapted from C. I methyl D tryptophan (Ido inhibitor). Adapted from
Figure 3. An illustration of the crystal structure for Indoleamine 2,3-dioxygenase (IDO) and chemical structures of its Inhibitors. A. bound to an inhibitor molecule.Indoleamine-2,3-dioxygenase inhibitors. Adapted from B. INCB024360 adapted from C. I methyl D tryptophan (Ido inhibitor). Adapted from

Novel Therapeutic Approaches to Leishmania Infection

Levi H.C. Makala1 and Babak Baban2

1. Introduction

1.1. Leishmaniasis

Leishmaniasis is a parasitic disease transmitted by phlebotomine sandflies. Approximately 1.2 million cases of cutaneous leishmaniasis (CL) and 500,000 cases of visceral leishmaniasis (VL), which is lethal if untreated, occur annually across the globe as per world health organization (WHO) estimates [1-3]. Current statistics and information relevant to leishmaniasis are summarized in Table 1. Leishmaniasis currently affects about 12 million people and it is estimated that approximately 350 million people live in risk of infection [1-3].The number of cases of leishmaniasis is probably underestimated because only 40 of the 88 countries where diseases frequently occur report them on a regular basis [4]. Leishmaniasis, is caused by several leishmania spp., that are obligate intracellular and unicellular kinetoplastid protozoan flagellate that establish themselves within the phagolysosome of host immune competent cells, especially macrophages and dendritic cells (DCs). In 1903, W.B. Leishman and C. Donovan reported this new parasite at the turn of the century [5,6]. Ronald Ross christened the new genus leishmania and the new species donovani in year 1903 [7]. L. major infection (leishmaniasis) in mice is a widely used model of human infection that has yielded critical insights into the immunobiology of leishmaniasis [8-10]. Leishmaniasis as a parasitic disease manifests itself mainly in 3 clinical forms; visceral leishmaniasis (VL), cutaneous leishmaniasis (CL) and mucocutaneous leishmaniasis (MCL), of which VL is the most severe form of the disease. VL is lethal if untreated and spontaneous cure is extremely rare. Cutaneous leishmaniasis usually has milder course and often results into a self-healing of ulcers. Resolution of leishmanial infection is dependent on the coordinated interactions between components of cell mediated immune response, specifically the activation of targeted T-cell populations for appropriate cytokine production and activation of macrophages. L. major infection of B6 and BALB/c mouse strains drives predominantly TH1 and TH2 responses, respectively [11-14]. In murine model, the development of Th1 response is associated with control of infection, and Th2 response is associated with disease progression. However, Th1 and Th2 dichotomy in the human system is not as distinct as in mice and the murine model does not strictly apply to human leishmaniasis.

Parameter Statistic or Information
Geographical locationWorldwide tropical and subtropical regions
Population at risk in 2013~350 million
Number of people affected~12 million
Number of deaths in 2013~20,000 – 30,000
Number of new cases in 2013~1.3 million
Global disease burden in 2013 (DALYs)~1.7 million
Multidrug-resistance in 2013Resistance to antimonials only
Visceral Leishmaniasis (VL)~200 000 to 400 000 new cases of VL occur worldwide each year. Over 90% of new cases occur in six countries: Bangladesh, Brazil, Ethiopia, India, South Sudan, and Sudan.
Cutaneous Leishmaniasis (CL)~One-third of CL cases occur in the Americas, the Mediterranean basin, and the Middle East and Central Asia. An estimated 0.7 million to 1.3 million new cases occur worldwide annually
Mucocutaneous LeishmaniasisReported in Bolivia, Brazil and Peru.
Major risk factorsSocioeconomic conditions, Malnutrition, Population mobility, Environmental changes, Climate change
Prevention and controlEarly diagnosis and effective case management, Vector control, Effective disease surveillance, Control of reservoir hosts, Social mobilization and strengthening partnerships

Table 1.

Factfile: WHO leishmaniasis statistics for 2013 (Adapted from

[i] - Abbreviations: CL, cutaneous leishmaniasis; DALYs, disability-adjusted life years; NK, not known; VL, visceral leishmaniasis, WHO, World Health Organization.

2. Conventional treatment strategies and limitations

Chemotherapy is the primary method used to control leishmaniasis. Despite the existence of several drugs for chemotherapy of human leishmaniasis, many of them are new formulations of ancient drugs repurposed in the last decade [15,16]. The treatment options for leishmaniasis are limited and include penta-valent antimonials, pentamidine, amphotericin B (AmB) and its lipoidal formulations and miltefosine, which have been introduced recently in the group of antileishmanial drugs (Table 2). Among all of these drugs, pentavalent antimonials are the first choice drugs in most of the developing countries as in these countries treatment strategy is governed by economic factors. But a large number of incidences of resistance have been observed for antimonials, particularly in India where the failure rate has been reported up to 65% [17,18]. AmB is very effective against leishmania parasite but frequent and severe adverse effects associated with it limit its application.

Drugs Admin Route Dosage Known Toxicities Mechanism of Action Resistance Comment References
Antimonial drugs (sodium stibogluconate and meglumine antimoniate (Pentostam)IM, IV28 mg/kg/day (28-30 days)Cardioto-xicity, nephroto-xicity, hepatotoxicity, pancreati-tis (frequent and severe) Activated within the amastigote, but not in the promastigote, by conversion to a lethal trivalent form. Activation. Mechanism not known. Antileishmanial activity might be due to action on host macrophage.Failure rates up to 65% (in India)First line drugs but high incidences of resistance has been emerged[16,19-22]
Amphotericin B (AmB) or Polyene antibioticIV0.75-1 mg/kg/day (15-20 days, daily or alternately)Severe nephrotoxicity, infusion-related reactions (frequent and severe) Complexes with 24-substituted sterols, such as ergosterol in cell
membrane, thus causing pores which alter ion balance and result in cell
- Severe toxicity[16,22,23]
Lipoidal formulations of AmB (Amphotec or Amphocil; AmBisome; Abelcet and dimyristoyl phosphatidyl glycerol with AMB)IV10-30 mg/kg total dose (single dose 3-5 mg/kg/dose)Mild nephrotoxicity (infrequent and mild)AmB formulation, act by binding to the sterol component cell membranes, leading to alterations in cell permeability and cell death. They bind to the cholesterol component of the mammalian cell. - High market price[16,22,24,25]
Miltefosine (Hexadecylphosphocholine)Oral100 mg/day (28 days)GIT problems, nephrotoxicity, hepatotoxicity, chances of teratogenicity (frequent, mild and transient)Primary effect uncertain, possible inhibition of ether remodelling,
phosphatidylcholine biosynthesis, signal transduction and calcium
Common in laboratory isolates Effective orally but its long half-life may encourage emergence of resistance on prolonged use[16,22,26,27]
Paromomycin (Monomycin or AminosidineIM15 mg/day (21 days)Nephrotoxicit- ototoxicity and hepatotoxicity (infrequent)In bacteria, paromomycin inhibits protein synthesis by binding to 30S subunit ribosomes, causing misreading and premature termination of
mRNA translation. In Leishmania, paromomycin also affects mitochondrion.
Common in laboratory isolatesLow cost; being investigated by non-profit groups[16,22,26]

Table 2.

Standard treatment protocols for leishmaniasis, characteristics and mechanisms of action

[i] - Adapted with modifications from Jain and Jain, 2013 and van Griensven, J. and Diro, E. 2012)

[ii] - Abbreviations: AmB, Amphotericin B; IM, Intramascular; IV, Intravenous;

The development of lipoidal formulations of AmB reduced the severity and frequency of adverse effects but resulted in high cost of formulation [22-25]. The conventional treatment schedule for visceral leishmaniasis suffers from a lot of problems like invasive route of administration (parenteral), long treatment course, severe toxicity (nephrotoxicity, cardiotoxicity, among others), high cost of treatment, few treatment regimens, emergence of resistance and variable patient response [17,28,29]. Thus there is continuous need for alternative new treatment strategies, vaccine candidates and new chemotherapeutic agents to provide complete cure from leishmaniasis taking into account the fatality of disease, high toxicity, high cost and inefficiency of current treatment protocols.

2.1. Nano-based antileishmanial agents

Currently, the pharmaceutical industry has undergone a profound transformation with the advent of nano-science. With a rapid growth of nanotechnology, different nanoparticles have been presented for medical science applications. Nanomaterials have unique chemical and physical properties, and may be used in the treatment of different severe or chronic diseases in the future [30]. Hitherto, it has been shown that some metal and metal oxide nanoparticles have antimicrobial activities [31]. It has long been demonstrated that silver ions, silver nanoparticles (Ag NPs), and nanosilver-containing complexes have antimicrobial behavior with high ability to inactivate bacteria and viruses [32]. Other reports indicate that gold nanoparticles (Au NPs), titanium dioxide nanoparticles (TiO2 NPs), zinc oxide nanoparticles (ZnO NPs), magnesium oxide nanoparticles (MgO NPs), etc. have antibacterial properties [33-37]. Nanotechnology has enabled the creation of nano-particle formulations such as liposomes, microemulsions and microcapsules [34]. Liposomes are microscopic vesicles composed of one or more concentric lipid bilayers separated by aqueous media. They can encapsulate hydrophilic and lipophilic substances in the aqueous compartment of the membrane respectively. Since liposomes are biodegradable, biocompatible and non-immunogenic, they are highly versatile for research, therapeutic and analytical applications [38]. In spite of the reported antileishmanial properties of nanoparticles under UV, IR, and dark conditions, these nanoparticles have the some cytotoxicity on immune competent cells such as macrophages and other antigen presenting cells and this must be considered in future applications and studies.

2.2. Leishmaniasis vaccines

Despite the knowledge about various life stages of the parasite and the ongoing work, designing an effective vaccine against leishmaniasis is still a matter of research, there are hundreds of potent vaccine candidates but issues regarding the cost, antigenic complexity along with the variability of organisms and the mixed type of responses produced in the host are limiting the progress in the relevant direction. Thus the technical challenges and the complexity in the immunity against the parasites clearly contribute to the absence of vaccines. There are three vaccines known: two in Brazil and one in Europe out of which one is highly efficient in treating VL and CL, thus still enlightening the ray of hope for progress in this field [39]. A glimpse of various antigens that has been used as vaccine candidates in last two decades are summarized in (Figure 1).


Figure 1.

A glimpse of various antigens that have been used as vaccine candidates in past ten years (Adapted and modified with permission from Singh, and Sundar, 2012).

These candidates include major surface, intracellular, stress responsive molecules, as well as other biomolecules of various metabolic pathways that can be the targets for vaccine development. Vaccine design and development has focused on all the forms of leishmania because of the conserved nature of molecules in all the species of leishmania that have been selected as the targets. Many of these targets have been studied in mice models while others in humans during diseased state producing promising results (Table 3). The availability of complete genome sequence of leishmania has provided hope for researchers to work with novel molecules as vaccine candidates [40]. The history of vaccination with the virulent forms of leishmania termed as “leishmanization” dates back to early 20th century but has since been banned for trials due safety concerns in human models [41]. First generation vaccines were limited in terms of the conferred immunity [42]. Second generation vaccines are currently in trial and are useful in providing protection of varying levels in different species along with the DNA and other subunit vaccines. The main hurdle in developing a potent vaccine stems from lack of multiple experimental study models necessary to provide all facets of immune responses in humans as well as safety issues [43].

Antigenic Molecule Vaccine type Experimental model References
LPG, gp 63, A2Native antigen Mouse[ 44-47]
gp 63Protein expressed in BCGMouse[48]
gp63, gp 46, PSA-2, A2, KMP11DNA VaccineMouse[49-53]
p36, LACKDNA vaccine + protein expressed in vaccinia virusMouse[54]
p36, LACKRecombinant protein + IL12Mouse[55,56]
LmSTI1, TSA, HASPB1, CPBRecombinant proteinMouse[57-59]
gp 63, KMP11DC pulsed with native antigenMouse and Monkey[60]
gp 63, LCR 1Protein expressed in BCGMouse[61,62]
Leish 111fRecombinant polyprotein of TSA, LmSTI1
and LeIF + MPL-SE
IDO 1IDO Inhibitors / IDO specific vaccinesMouse and Human[65-68]

Table 3.

Current Potent/effective vaccines against leishmaniasis

[i] - DC – dendritic cells CP – cysteine proteinase; BCG Mycobacterium bovis bacillus Calmette–Guerin; IDO, Indoleamine pyrrole 2,3-dioxygenase, IL, interleukin; MPL-SE, monophosphoryl lipid A soluble emulsion; TSA, thiol-specific-antioxidant antigen.

2.3. Potential drug targets

Notwithstanding the significant progress of leishmanial research in the last few decades, identification and characterization of novel drugs and drug targets are far from satisfactory. The digenetic life cycle of leishmania consists of motile flagellated, extracellular promastigote forms which survive and multiply within the phagolysosomal compartments of macrophages and other antigen presenting cells. Therefore, the search for new potential drug targets mainly focuses on biochemical and metabolic pathways essential for parasite survival [69-71]. The strategy to target more than one enzyme of a metabolic pathway simultaneously may prove more useful and effective. Important biochemical and enzymatic machineries that are utilized as putative drug targets for generations of true antileishmanial drugs are as follows: enzymes of polyamine synthesis [72,73], enzymes of the glycosomal machinery [74,75], enzymes of thiol-metabolic cyclin dependent kinases, enzymes of sterol biosynthesis [76,77], Pepsidases, Mitogen activated protein kinases (MAPK), dihydrofolate reductases (DHFR) [78], topoisomerases metacaspases [79,80]; and leishmanial antigens that modulate host immune functions [81].

Polyamines are not only involved in parasite growth and differentiation, but also down regulate lipid peroxidation generated by oxidant compounds to make the environment compatible for survival [82-84]. The leishmania genome has 154 peptidases namely serine, cysteine, aspartic, threonine and metallopeptidases. These enzymes play a role in reducing viability and induction of morphological changes [85,86]. Roles of other enzyme systems include but limited to roles in metabolic activities like glycolysis, oxidation of fatty acids, lipid biosynthesis and purine salvage pathways [87-103]. Other Functions of the above mentioned biochemical and metabolic pathways include: Cell division cycle, transcription, apoptosis, cell proliferation, cell differentiation and innate immunity to activation of adaptive immunity [104-113]. All these functions are essential for parasite survival, hence can be targeted to disrupt the unique targeting signal sequences.

3. Phytotherapy

Phytotherapy is the study of the use of extracts from natural origin as medicines or health-promoting agents. The main difference of phytotherapy medicines from the medicines containing the herbal elements is in the methods of plant processing. Methods of plant processing to receive medicines containing herbal elements are aimed on extraction of the chemical clean active substances, but methods of plants processing to obtain phytotherapy medicines are aimed to preserve all complex of active substances of plant in the most simple and close to the natural form. The biological activity of plant extracts has been attributed to compounds belonging to diverse chemical groups including alkaloids, flavonoids, phenylpropanoids, steroids, and terpenoids. [114-116]. Phytotherapy can be an important tool in the search for novel antileishmanial agents with fewer side effects and low cost. Firstly, the chemical diversity of plants makes them a valuable source of metabolites with pharmacological relevance [117]. Secondly, metabolites isolated from plants extracts or essential oils can be used in several different ways in the development of drugs. To obtain a herbal medicine or an isolated active compound, different research strategies can be employed, among them, investigation of the traditional use, the chemical composition, the toxicity of the plants, or the combination of several criteria [118]. In the extraction processes, different plant parts and different solvents have generally been used. In screening for biological activity, there is clearly substantial room for improvement in the extraction methodologies, since a variety of techniques can be used to prepare extracts [119-121]. Usually, solvents of different polarity are employed for the extractions. For purification and isolation, the active extracts of the plant are sequentially fractionated, and each fraction and/or pure compound can be evaluated for biological activity and toxicity. This strategy is called bioactivity-guided fractionation, which allows tests that are simple, reproducible, rapid, and low-cost [120,121]. In the last two decade, much attention has been given to the search of novel drug delivery systems for herbal drugs. The development of nanoparticles loaded with herbal drugs presents several advantages including: increase of drug solubility and bioavailability, protection against the toxicity, enhancement of pharmacological activity, increase of stability, and protection against degradation [122]. The tendency of nanoparticules, especially liposomes, to be captured by the mononuclear phagocyte system may be an additional advantage in the treatment of a variety of intracellular infectious diseases. Intraperitoneal and intravenous administration of liposomes proved to be a good bio-distribution system for drugs in the treatment of visceral leishmaniasis, since it allows increasing of drug accumulation in macrophage-rich tissues such as liver and spleen thus reducing the level of toxicity to other tissues and organs [123].

In vitro screenings are only the first steps to prove the efficacy and safety of medicinal plants for application in the treatment of leishmaniasis. In addition, variation in the efficacy of drugs in treating leishmaniasis may often result from differences in the drug sensitivity of leishmania species, the immune status of the patient, or the pharmacokinetic properties of the drug [4]. A review of the literature on the use of natural products, including plant crude extracts, fractions, isolated compounds, and essential oils, shows that there is a massive effort by scientists around the world to identify and characterize natural plant compounds with antileishmanial activity [124-126]. These efforts are now bearing fruit, obtaining good results and validating natural products as genuine sources for drug discovery. A fitting example would be essential oils that are known to possess a wide variety of hydrophobic compounds with antimicrobial potential. The ability to diffuse across cell membranes certainly gives to those molecules some advantage in targeting cellular components, being a valuable research option for the search of bioactive compounds [124-126]. The Ocimum gratissimum essential oil and eugenol, its major component, was tested on the growth, viability, and ultrastructural alterations of the amastigote and promastigote forms of L. amazonensis, as well as on the interaction of these flagellates with mouse peritoneal macrophages, concomitant with nitric oxide production stimulation by the infected macrophages. Significant mitochondrial alterations occurred at the ultrastructural level of the parasite, such as remarkable swelling, disorganization of the inner membrane, and an increase in the number of cristae after treatment of parasites with O. gratissimum essential oil [125, 126]. Additionally, the linalool-rich essential oil extracted from the leaves of Crotoncajucara, also has effects on L. amazonensis parasites, on the interaction of these flagellates with mouse peritoneal macrophages and on nitric oxide production by the infected macrophages [125].

3.1. Alternative synthetic compounds

Screenings of synthetic compounds and their derivatives for antileishmanial activity has been carried out. In the last 10 years several compounds have been tested to identify potential new drugs, with the desirable characteristics. Most of the compounds exhibited antileishmanial activity against the promastigote form of L. major at non-cytotoxic concentrations and these compounds are also effective against intracellular L. major, and significantly decrease the infectivity index [127,128].

The stilbene trans-3,4 0,5-trimethoxy-3 0 -amino-stilbene (TTAS) has potent effect with low toxicity on Leishmania infantum (LD 50 value of 2.6 g/mL). The mechanism of action involves the disruption of the mitochondrial membrane potential and the ability to block leishmania parasites in the G2–M phase of the cell cycle [129,130]. N -Butyl-1-(4-dimethylamino) phenyl-1,2,3,4- tetrahydro- β -carboline-3-carboxamide is effective against Leishmania amazonensis.

BTB 06237 (2-[(2,4-dichloro-5-methylphenyl)sulfanyl]-1,3-dinitro-5-(trifluoromethyl) benzene) and its analogues, a compound previously identified through quantitative structure-activity relationship (QSAR) has also been shown, to possesses potent and selective activity against leishmania parasites. This compound and its analogues has the ability to reduce parasitemia levels in immune competent cells especially peritoneal macrophages, and additionally possess the ability to generate reactive oxygen species (ROS) in L. donovani promastigotes [131]. The in vitro antileishmanial activity of 44 derivatives of 1,3,4-thiadiazole and related compounds against promastigote forms of L. donovani have also been tested. Micromolar concentrations of these agents were used to study the inhibition of multiplication of promastigotes. Seven compounds were identified as potential antigrowth agents against the parasite [132]. Additionally, a series of 2,4,6-trisubstituted pyrimidines and 1,3,5-triazines following synthesis and screening display antileishmanial activity against L. donovani. Nitroimidazolyl-1,3,4-thiadiazole-based antileishmanial agents against L. major also exhibits antileishmanial activity against the promastigote form of L. major at non-cytotoxic concentrations [128]. Other compounds with antileishmanial activity, both in vitro and in vivo include a series of 1- phenylsubstituted b-carbolines containing an N-butylcarboxamide group at C-3 of the b-carboline nucleus, tetrahydrobenzothieno-pyrimidines, R(+)-limonene derivatives, quinoline tripartite hybrids from chloroquine, ethambutol, and isoxyl drugs, and (4-butoxyphenyl)-N0-{2-[(7-chloroquinolin-4-yl)amino]ethyl}urea [133,136]. The urgent need to develop cost-effective new drugs and to discover novel molecules with potent antiparasitic activity and improved pharmacological characteristics cannot be overemphasized. Although many advances have been made in the treatment of leishmaniasis, much still remains to be understood.

3.2. A potential role of indoleamine 2,3-dioxygenase-specific T cells in leishmania vaccination

IDO is an immunoregulatory enzyme implicated in immunity under normal and pathological settings [137,138], and provides a potential mechanism for the development of dendritic cell (DC)–mediated T-cell tolerance [139]. IDO1 DCs inhibit T-cell proliferation due to tryptophan depletion and accumulation of toxic tryptophan metabolites [138]. 1-Methyl-D-tryptophan is an inhibitor of the enzymes IDO and INDOL1 (indolamine 2,3-dioxygenase 1 and 2) with selectivity for INDOL1 [140-142]. The enzymes perform similar transformations and are responsible for catalyzing the rate-limiting step of oxidative tryptophan catabolism in the kynurenine pathway. IDO activity is correlated with an induction of tolerance and immune-suppression through activation of regulatory T cells by metabolites generated from tryptophan catabolism. Inhibition of IDO by 1-Methyl-D-tryptophan blocks this induced immune suppression, which has shown utility in suppressing acquired immunities of tumors and indicates potential for chemical intervention of chronic inflammatory diseases [143-145]. In a recent report, Makala and colleagues [65, 66] elegantly showed that IDO is implicated in suppressing T-cell immunity to parasite antigens, and IDO inhibition reduced local inflammation and parasite burdens. The findings by Makala and colleagues support a counter-regulatory role for IDO that benefits the pathogen, not the host. In this regard, an interesting aspect of IDO is that systemic inactivation at the organism level, either pharmacologically or genetically, does not appear to cause autoimmunity [65-68,138]. A conceptual model of IDO-mediated activation and effector T cell suppression following L. major infection is summarized in Figure 2 [Makala, 2012].


Figure 2.

A conceptual model of IDO-mediated activation and effector T cell suppression following L. major infection. The model depicts interactions between IDO+ DCs, Tregs and naïve T cells that drive suppressive and non-suppressive outcomes under IDO-sufficient (+) and IDO-deficient (-) conditions in response to L. major infection. Induced IDO activity in DCs triggers cell stress responses blocks IL6 production by pDCs themselves, and by other cells (e.g. macrophages) capable of producing IL6. Under conditions of IDO ablation the same stimuli do not create suppression, and instead DCs stimulate naïve T cells, and express IL6, which converts Tregs to TH17 T cells or promotes TH17 differentiation from naïve CD4+ T cells (Adapted and modified from Makala, 2012).

The model depicts interactions between IDO+ DCs, Tregs and naïve T cells that drive suppressive and non-suppressive outcomes under IDO-sufficient (+) and IDO-deficient (-) conditions in response to L. major infection. Induced IDO activity in DCs triggers cell stress responses blocks IL6 production by pDCs themselves, and by other cells (e.g. macrophages) capable of producing IL6. Under conditions of IDO ablation the same stimuli do not create suppression, and instead DCs stimulate naïve T cells, and express IL6, which converts Tregs to TH17 T cells or promotes TH17 differentiation from naïve CD4+ T cells. The chemical structures of IDO and its inhibitors are shown in Figure 3.


Figure 3.

An illustration of the crystal structure for Indoleamine 2,3-dioxygenase (IDO) and chemical structures of its Inhibitors. A. bound to an inhibitor molecule.Indoleamine-2,3-dioxygenase inhibitors. Adapted from B. INCB024360 adapted from C. I methyl D tryptophan (Ido inhibitor). Adapted from

To examine the possible effects (and/or side effects) of the induction of IDO-specific T cells, a phase I vaccination study is ongoing at the Center for Cancer Immune Therapy, Copenhagen University Hospital, in which patients with non-small-cell lung cancer are vaccinated with an IDO-derived peptide with Montanide adjuvant (; NCT01219348). Different species of leishmania are responsible for cutaneous, mucocutaneous, or visceral leishmaniasis infections in millions of people and animals. Adverse reactions caused by antileishmanial drugs, emergence of resistance, and lack of a vaccine have motivated the search for new therapeutic options to control this disease. There have been notable advances in molecular diagnostics, in the understanding of host immune responses to infection, and in vaccine development. The fact that IDO may be involved in tolerance to non-self-antigens, may have key attractive implications for IDO-based immune therapy as boosting immunity to neo-antigens, but not normal self-antigens, by the activation of IDO-specific T cells. Makala and colleagues [65-68, 138] demonstrated that IDO suppresses adaptive immunity, supporting the notion that in clinical setting, the targeting of IDO could have synergistic effects in leishmania vaccine development. Thus, the induction of IDO-specific immune responses by therapeutic measures could function synergistically with additional immune therapy. Almost any successful immune therapy strategy aims at inducing immunological activation and inflammation. Since IDO-expressing cells might antagonize the desired effects of other immunotherapeutic approaches, targeting IDO-expressing cells by vaccination would be easily implementable and compatible with such therapeutic measures.

3.3. Multidrug treatment strategy

Combination therapy has increasingly been explored, particularly in highly endemic regions, aiming to identify a short, cheap, well-tolerated combination regimen that can preferably be given in an ambulatory way and requiring minimal clinical monitoring. To date combination therapy has shown promising results including improved treatment efficacy with reduced side effects, shorter duration of therapy, reduced cost as well as reduced incidence of resistance in phase 2 clinical trial, as has been used for diseases like malaria, tuberculosis, and HIV. [1,3,146]. A 17-day combination of antimonials with paromomycin was found effective in east Africa (93% efficacy). Owing to such fascinating results numerous phase 3 clinical trials are progressively being conducted in Asian and African continents to further investigate the clinical efficacy of combination therapy in treatment of leishmaniasis. Researchers have continued to study the effect of immunotherapy using combinations of two or more antileishmanial drugs [146-149]. A list of completed or currently in progress clinical trials for treatment of leishmaniasis are shown in (Table 4). Sundar et al. [150] investigated the efficacy and safety of three combinations of three effective antileishmanial drugs (lipo-somal AmB, miltefosine, paromomycin) and compared their efficacy as duration of treatment with the standard monotherapy in India, that is, AmB infusion in an open-label, parallel-group, non-inferiority, randomized control trial conducted in two hospitals at Bihar, India. Combination regimens including liposomal amphotericin B (5 mg/kg single dose), paromomycin and/or miltefosine were also found highly effective (98%–99%) and safe, and are now included in WHO recommendations for the Indian subcontinent [3,146,147]. The multidrug treatment has been found equally effective as standard monotherapy even with fewer side effects and shorter course of administration [150]. Combination treatment approaches for leishmaniasis have been advocated by many scientists but they are also enforcing the simultaneous development of other measures in the control of this parasitic disease in the endemic regions of Asia and Africa including control of sandflies, clinical monitoring of treatment, advances in case detection and rapid methods of diagnosis as well as proper evaluation of various leishmania control programs [146,151,152]. The clinical efficacy of multidrug therapy has been confirmed and so far the results are convincing and give hope for the future in terms of treatment.

Condition Intervention Study Phase Study Objective
• Meglumine antimoniate (MA)2
To compare efficacy of the standard recommended schedule with an alternative regimen of MA in the treatment of ML /MCL
CL• Paramomycin
• WR279, 396 ( Paramomycin / Gentamycin)
3To determine if WR279, 396 results in statistically superior final clinical cure rates compared to Paramomycin alone
VL• Antimoniate of N-Methylglucamine (Fungizone)
• Amphotericin B Deoxycholade (Anforicin)
• Liposomal Amphotericin B (Ambisome)
4To compare efficacy and safety of medications in Brazil
L• Sodium Stibogluconate (SSG) (Pentosam)2To collect safety and efficacy data on the use of Pentosam
VL• AmBisome + Miltefosine
• AmBisome + Paromomycin sulfate
• Miltefosine + Paromomycin sulfate
• Amphotericin B Deoxycholate
3To identify a safe and effective combination for short course treatment of visceral leishmaniasis with reduced risk of parasite resistance
VL• Sodium stibogluconate
• Paromomycin sulfate
• Sodium stibogluconate + Paromomycin sulfate
3To assess the efficacy and safety of sodium stibogluconate 30 days alone, paromomycin sulfate 21 days alone and sodium stibogluconate and paromomycin sulfate as a combination course of 17 days in the treatment of patients with visceral leishmaniasis
VL• Ambisome
• AmBisome + Miltefosine
• AmBisome + Paromomycin
• Miltefosine + Paromomycin
3To evaluate efficacy and safety of various combinations of the three drugs; AmBisome, paromomycin and miltefosine at reduced total dosage against the standard treatment with a total dose of 15 mg/kg of AmBisome
VL• AmBisome + Sodium stibogluconate
• AmBisome + Miltefosine
• Miltefosine
2To assess combinations of sodium stibogluconate plus single dose AmBisomeW, miltefosine plus single dose AmBisomeW and miltefosine alone in treatment of visceral leishmaniasis in Eastern Africa
VL• Miltefosine + AmBisome2To sequential design to combine miltefosine and AmBisome in different doses
VL• AmBisome + Miltefosine2To evaluate the final cure after six months on sequential administration of both drugs. AmBisome will be given on day 1, followed by miltefosine for 14 days
VL• Sitamaquine2To evaluate the final cure after six months on sequential administration of both drugs. AmBisome will be given on day 1, followed by miltefosine for 14 days

Table 4.

Clinical trial completed or currently recruiting for treatment of leishmaniasis (at: (accessed 10-10-2013))

[i] - Mucosal Leishmaniasis (LM), Mucocutaneous Leishmaniasis (MCL), Cutaneous Leishmaniasis (CL), Visceral Leishmaniasis (VL), Meglumine antimoniate (MA), Leishmaniasis (L).

4. Concluding remarks

Leishmaniasis is one of the major neglected infectious diseases. Progress has been achieved in terms of treatment, including the development of combination therapy as well as our understanding of the molecular nature of potential vaccine candidates following the completion of the genome sequence. The occurrence of drug resistance in disease-endemic countries is concerning and should be closely monitored. In spite of all these drawbacks, there is presently rapid progress in our understanding of the molecular nature of potential vaccine candidates. There is a need to develop more potent, cost effective drugs and vaccine candidates. Total eradication of leishmaniasis will depend on the combined efforts of governments, the scientific research community, the pharmaceutical industry and people with a view to reduce the transmission of disease, rapid diagnosis and appropriately targeted treatment of the various forms of leishmanisis. Understanding of the molecular nature of potential vaccine candidates could potentially lead to novel gene-based, plant-based and synthetic-based therapeutic approaches or a dependable cure for leishmaniasis.


LM: Mucosal leishmaniasis

MCL: Mucocutaneous leishmaniasis

CL: Cutaneous leishmaniasis,

VL: Visceral leishmaniasis

MA: Meglumine antimoniate

AmB: Amphotericin B

IM: Intramascular

IV: Intravenous

DC: dendritic cells

CP: cysteine proteinase

BCG: Mycobacterium bovis bacillus Calmette–Guerin

IDO: Indoleamine-pyrrole 2,3-dioxygenase

IL: Interleukin

MPL-SE: monophosphoryl lipid A soluble emulsion

TSA: thiol-specific-antioxidant antigen

pDC: Plasmacytoid Dendritic Cell

HSP: Heat Shock Proteins

LmSTI1: L. major stress-inducible 1

LeIF: Leishmania elongation initiation factor

KMP-11: Kinetoplastid membrane protein-11

GSH: Glutathione complex

TSH: thyroid - stimulating hormone

LACK: Leishmania analogue of the receptor kinase C

Lcr1: T-cell antigens from an amastigote of L. chagasi containing homologous 67-amino-acid repeats

Ldp23: 23 kDa highly hydrophilic protein rich in lysine residues present on the surface of L. donovani and L. major

LPG: Leishmania major lipophosphoglycan

T(SH)2: trypanothione;

CRK: cdc-2 related kinase

RIC: RNA import complex

A2: amastigote stage-specific protein family in L. donovani

HASPB: hydrophilic acylated surface protein B

PFR-2paraflagellar rod protein

MAPK: Mitogen-activated Protein (MAP) kinases

SMT: sterol 24-cmethyltransferase

GPI/GP46: glycosylphosphatidylinositol

PSA: Promastigote surface antigen

MML: multi-subunit recombinant leishmanial vaccine


1 - World Health Organization. First WHO report on neglected tropical diseases. 2012. WHO. 1-172. Available at:
2 - Aguilar-Be I Zardo RS, Souza EP, Borja-Cabrera GP, Rosado-Vallado M, Mut-Martin M, García-Miss MR, Sousa CBP, Dumonteil E. Cross-Protective Efficacy of a Prophylactic Leishmania donovani DNA Vaccine against Visceral and Cutaneous Murine Leishmaniasis. Infection and Immunity. 2005;73(2) 812–819.
3 - World Health Organization. Control of the leishmaniasis. Report of a meeting of the WHO Expert Committee on the Control of Leishmaniases. Geneva (Switzerland): World Health Organization; 2010. Available at: (Accessed October 10, 2013).
4 - Croft SL, Sundar S, Fairlamb AH. Drug resistance in leishmaniasis. Clinical Microbiology Review 2006;19(1) 111–126.
5 - Leishman W.B: "On the possibility of the occurrence of trypanomiasis in India". The British Medical Journal 1903
6 - Donovan C: "Memoranda: On the possibility of the occurence of trypanomiasis in India.".The British Medical Journal 1903.
7 - Ross R: "Further notes on Leishman's bodies". Ibid 1903;(ii) 1401.
8 - Peters N, Sacks D. Immune privilege in sites of chronic infection: Leishmania and regulatory T cells. Immunological Reviews 2006;213 159–179.
9 - Liese J, Schleicher U, Bogdan C: The innate immune response against Leishmania parasites. Immunobiology 2008;213 377–387.
10 - Sharma U, Singh S: Immunobiology of leishmaniasis. Indian Journal of Experimental Biology 2009;47 412– 423. 161.
11 - Moll H, Röllinghoff M. Resistance to murine cutaneous leishmaniasis is mediated by TH1 cells, but disease-promoting CD4+ cells are different from TH2 cells. European Journal of Immunology 1990;20(9) 2067-2074.
12 - Lohoff M, Sommer F, Solbach W, Röllinghoff M. Coexistence of antigen-specific TH1 and TH2 cells in genetically susceptible BALB/c mice infected with Leishmania major. Immunobiology. 1989;179(4-5) 412-421.
13 - Locksley RM, Heinzel FP, Holaday BJ, Mutha SS, Reiner SL, Sadick MD. Induction of Th1 and Th2 CD4+ subsets during murine Leishmania major infection. Research in Immunology 1991;142(1) 28-32.
14 - Scott P. IFN-gamma modulates the early development of Th1 and Th2 responses in a murine model of cutaneous leishmaniasis. Journal of Immunology 1991;147(9) 3149-3155.
15 - Croft SL, Yardley V. Chemotherapy of leishmaniasis. Current Pharmaceutical Design 2002;8 319-342.
16 - Jain K, Jain NK. Novel therapeutic strategies for treatment of visceral leishmaniasis. Drug Discovery Today 2013 [Epub ahead of print].
17 - Sane, S.A. Shakya N, Gupta S. Immunomodulatory effect of picroliv on the efficacy of paromomycin and miltefosine in combination in experimental visceral leishmaniasis. Experimental Parasitology 2011;127(2) 376–381.
18 - Ait-Oudhia K. Gazanion E, Oury B, Vergnes B, Sereno D. The fitness of antimony-resistant Leishmania parasites: lessons from the field. Trends in Parasitology 2011;27 141–142.
19 - Roberts WL, McMurray WJ, Rainey PM. Characterization of the antimonial antileishmanial agent meglumine antimonate (glucantime). Antimicrobiol Agents of Chemotherapy 1998;42(5) 1076-1082.
20 - Zilberstein,D. and Ephros, M. (2002) Clinical and laboratory aspects of Leishmania chemotherapy in the area of drug resistance. In World Class Parasites (Vol. 4) (Black, S.J. and Seed, J.R., eds), pp. 115–136, Kluwer Academic Press, London.
21 - Pathak, M.K. and Yi, T. (2001) Sodium stibogluconate is a potent inhibitor of protein tyrosine phosphatases and augments cytokine responses in hemopoietic cell lines. J. Immunol. 167, 3391–3397.
22 - Croft SL, Coombs GH. Leishmaniasis--current chemotherapy and recent advances in the search for novel drugs. Trends in Parasitology 2003;19(11) 502-508.
23 - Roberts CW, McLeod R, Rice DW, Ginger M, Chance ML, Goad LJ. Fatty acid and sterol metabolism: potential antimicrobial targets in apicomplexan and trypanosomatid parasitic protozoa. Molecular Biochemical Parasitology 2003;126(2) 129-142.
24 - Clemons KV, and Stevens DA Comparative efficacies of four amphotericin B formulations--Fungizone, amphotec (Amphocil), AmBisome, and Abelcet--against systemic murine aspergillosis. Clemons KV, Stevens DA. Antimicrobiol Agents of Chemotherapy 2004;48(3) 1047-50.
25 - Daftarian PM, Stone GW, Kovalski L, Kumar M, Vosoughi A, Urbieta M, Blackwelder P, Dikici E, Serafini P, Duffort S, Boodoo R, Rodríguez-Cortés A, Lemmon V, Deo S, Alberola J, Perez VL, Daunert S, Ager AL. A Targeted and Adjuvanted Nanocarrier Lowers the Effective Dose of Liposomal Amphotericin B and Enhances Adaptive Immunity in Murine Cutaneous Leishmaniasis. Journal of Infectious Diseases 2013 [Epub ahead of print].
26 - el-On J, Halevy S, Grunwald MH, Weinrauch L. Topical treatment of Old World cutaneous leishmaniasis caused by Leishmania major: a double-blind control study. Journal of the American Academy of Dermatology 1992;27(2 Pt 1) 227-231.
27 - Croft SL, Seifert K, Duchêne M. Antiprotozoal activities of phospholipid analogues. Molecular Biochemical Parasitology 2003;126(2) 165-172.
28 - Akendengue B, Ngou-Milama E, Laurens A, Hocquemiller R. Recent Advances in the fight against leishmaniasis with natural products. Parasite 1999;6 3-8.
29 - Singh S.K. Bimal S, Narayan S, Jee C, Bimal D, Das P, Bimal R.. Leishmania donovani: assessment of leishmanicidal effects of herbal extracts obtained from plants in the visceral leishmaniasis endemic area of Bihar, India. Experimental Parasitology 2011;127(2) 552–558.
30 - Angeli E, Buzio R, Firpo G. Nanotechnology applications in medicine. Tumori 2008;94 206–215.
31 - Elechiguerra J.L, Burt JL, Morone, JR. Interaction of silver nanoparticles with HIV-1. Journal of Nanobiotechnology 2005;3 6.
32 - Shrivastava S, Bera T, Roy A, Singh G, Ramachandrarao P, Dash D. Characterization of enhanced antibacterial effects of novel silver nanoparticles. Nanotechnology 2007;18 1–9.
33 - AshaRani PV, Low Kah Mun G, Hande MP. Cytotoxicity and genotoxicity of silver nanoparticles in human cells. ACS Nano 2009;3 279–290.
34 - Chen M, Yang Z, Wu H, Pan X, Xie X, Wu C. Antimicrobial activity and the mechanism of silver nanoparticle thermosensitive gel. International Journal of Nanomedicine 2011;6 2873–2877.
35 - Li WR, Xie X., Sh, QS, Zeng H., OU-Yan, YS, Chen YB. Antibacterial activity and mechanism of silver nanoparticles on Escherichia coli. Appl. Microbiolology and Biotechnology 2010;85 1115–1122.
36 - Xia T, Kovochich M, Brant J. Comparison of the abilities of ambient and manufactured nanoparticles to induce cellular toxicity according to an oxidative stress paradigm. Nano Letters 2006;6 1794–1807.
37 - Zheng J, Wu X, Wang M, Ran D, Xu W, Yang J. Study on the interaction between silver nanoparticles and nucleic acids in the presence of cetyltrimethylammonium bromide and its analytical application. Talanta 2008;74 526–532.
38 - Edwards KA, Baeumner AJ. Liposomes in analyses. Talanta. London, 2006;68(5) 1432-1441.
39 - Singh B, Sundar S. Leishmaniasis: vaccine candidates and perspectives. Vaccine, 2012 30(26) 3834-3842.
40 - Kumari S, Kumar A, Samant M, Singh N, Dube A. Discovery of novel vaccine candidates and drug targets against visceral leishmaniasis using proteomics and transcriptomics. Current Drug Targets 2008;9(11):938–947.
41 - Noazin S, Modabber F, Khamesipour A, Smith PG, Moulton LH, Nasseri K, et al. First generation leishmaniasis vaccines: a review of field efficacy trials. Vaccine 2008;26 (52) 6759–6767.
42 - Rafati S, Nakhaee A, Taheri T, Taslimi Y, Darabi H, Eravani D, et al. Protective vaccination against experimental canine visceral leishmaniasis using a combination of DNA and protein immunization with cysteine proteinases type I and II of L. infantum. Vaccine 2005;23(28) 3716–25.
43 - Okwor I, Liu D, Uzonna J. Qualitative differences in the early immune response to live and killed Leishmania major: implications for vaccination strategies against Leishmaniasis. Vaccine 2009;27(19) 2554–62.
44 - Mitchell GF, Handman E, Spithill TW. Examination of variables in the vaccination of mice against cutaneous leishmaniasis using living avirulent cloned lines and killed promastigotes of Leishmania major. International Journal of Parasitology 1985;15(6) 677–684.
45 - McConville MJ, Bacic A, Mitchell GF, Handman E. Lipophosphoglycan of Leishmania major that vaccinates against cutaneous leishmaniasis contains an alkylglycerophosphoinositol lipid anchor. Proceedings of the National Academy of Science U S A 1987;84(24) 8941–8945.
46 - Rivier D, Bovay P, Shah R, Didisheim S, Mauel J. Vaccination against Leishmania major in a CBA mouse model of infection: role of adjuvants and mechanism of protection. Parasite Immunology 1999;21(9) 461–473.
47 - Soong L, Duboise SM, Kima P, McMahon-Pratt D. Leishmania pifanoi amastigote antigens protect mice against cutaneous leishmaniasis. Infection and Immunity 1995;63(9) 3559–3566.
48 - Abdelhak S, Louzir H, Timm J, Blel L, Benlasfar Z, Lagranderie M, et al. Recombinant BCG expressing the leishmania surface antigen Gp63 induces protective immunity against Leishmania major infection in BALB/c mice. Microbiology 1995;141(Pt 7) 1585–1592.
49 - Xu D, McSorley SJ, Chatfield SN, Dougan G, Liew FY. Protection against Leishmania major infection in genetically susceptible BALB/c mice by gp63 delivered orally in attenuated Salmonella typhimurium (AroA- AroD). Immunology 1995;85(1) 1–7.
50 - Walker PS, Scharton-Kersten T, Rowton ED, Hengge U, Bouloc A, Udey MC, et al. Genetic immunization with glycoprotein 63 cDNA results in a helper T cell type 1 immune response and protection in a murine model of leishmaniasis. Human Gene Therapy 1998;9(13) 1899–1907.
51 - Ahmed SB, Bahloul C, Robbana C, Askri S, Dellagi K. A comparative evaluation of different DNA vaccine candidates against experimental murine leishmaniasis due to L. major. Vaccine 2004;22(13–14) 1631–1639.
52 - Ghosh A, Zhang WW, Matlashewski G. Immunization with A2 protein results in a mixed Th1/Th2 and a humoral response which protects mice against Leishmania donovani infections. Vaccine 2001;20(1–2) 59–66.
53 - Basu R, Bhaumik S, Basu JM, Naskar K, De T, Roy S. Kinetoplastid membrane protein-11 DNA vaccination induces complete protection against both pentavalent antimonial – sensitive and – resistant strains of Leishmania donovani that correlates with inducible nitric oxide synthase activity and IL – 4 generation: evidence for mixed Th1 – and Th2 – like responses in visceral leishmaniasis. Journal of Immunology 2005;174(11) 7160–7171.
54 - Dondji B, Perez-Jimenez E, Goldsmith-Pestana K, Esteban M, McMahon-Pratt D. Heterologous prime – boost vaccination with the LACK antigen protects against murine visceral leishmaniasis. Infection and Immunity 2005;73(8) 5286–5289.
55 - Mougneau E, Altare F, Wakil AE, Zheng S, Coppola T, Wang ZE, et al. Expression cloning of a protective Leishmania antigen. Science 1995;268(5210) 563–566.
56 - Gurunathan S, Prussin C, Sacks DL, Seder RA. Vaccine requirements for sustained cellular immunity to an intracellular parasitic infection. Nature Medicine 1998;4(12) 1409–1415.
57 - Campos-Neto A, Porrozzi R, Greeson K, Coler RN, Webb JR, Seiky YA, et al. Protection against cutaneous leishmaniasis induced by recombinant antigens in murine and nonhuman primate models of the human disease. Infection and Immunity 2001;69(6) 4103–4108.
58 - Stager S, Smith DF, Kaye PM. Immunization with a recombinant stage – regulated surface protein from Leishmania donovani induces protection against visceral leishmaniasis. Journal of Immunology 2000;165(12) 7064–7071.
59 - Rafati S, Kariminia A, Seyde-Eslami S, Narimani M, Taheri T, Lebbatard M. Recombinant cysteine proteinases – based vaccines against Leishmania major in BALB/c mice: the partial protection relies on interferon gamma producing CD8(+) T lymphocyte activation. Vaccine 2002;20(19–20) 2439–2447.
60 - Berberich C, Ramirez-Pineda JR, Hambrecht C, Alber G, Skeiky YA, Moll H. Dendritic cell (DC) – based protection against an intracellular pathogen is dependent upon DC – derived IL – 12 and can be induced by molecularly defined antigens. Journal of Immunology 2003;170(6) 3171–3179.
61 - Connell ND, Medina-Acosta E, McMaster WR, Bloom BR, Russell DG. Effective immunization against cutaneous leishmaniasis with recombinant bacille Calmette– Guerin expressing the Leishmania surface proteinase gp63. Proceedings of the National Academy of Science U S A 1993;90(24) 11473–11477.
62 - Streit JA, Recker TJ, Donelson JE, Wilson ME. BCG expressing LCR1 of Leishmania chagasi induces protective immunity in susceptible mice. Experimental Parasitology 2000;94(1) 33–41.
63 - Coler RN, Goto Y, Bogatzki L, Raman V, Reed SG. Leish-111f, a recombinant polyprotein vaccine that protects against visceral Leishmaniasis by elicitation of CD4+ T cells. Infection and Immunity2007;75 (9) 4648–4654.
64 - Chakravarty J, Kumar S, Trivedi S, Rai VK, Singh A, Ashman JA, et al. A clinical trial to evaluate the safety and immunogenicity of the LEISH-F1 + MPL-SE vaccine for use in the prevention of visceral leishmaniasis. Vaccine 2011;29(19) 3531–3537.
65 - Makala LH, Baban B, Lemos H, El-Awady AR, Chandler PR, Hou DY, Munn DH, Mellor AL. Leishmania major attenuates host immunity by stimulating local indoleamine 2,3-dioxygenase expression. Journal of Infectious Diseases 2011; 203:715–725.
66 - Makala LH. The role of indoleamine 2, 3 dioxygenase in regulating host immunity to leishmania infection. Journal of Biomedical Science 2012;(9)19:5.
67 - Thor-Straten P, Andersen MH. A potential role of indoleamine 2,3-dioxygenase-specific T cells in leishmania vaccination. (Comment on Leishmania major attenuates host immunity by stimulating local indoleamine 2,3-dioxygenase expression. Journal of Infectious Diseases 2011;204(3) 488-489. doi: 10.1093/infdis/jir274.
68 - Mellor AL, Munn DH. Creating immune privilege: active local suppression that benefits friends, but protects foes. Nature Reviews Immunology 2008; 8:74–80.
69 - Gil ES, Cunha LC, Paula JR, Bezerra JCB, Aguiar, FA. Leishmaniasis: Therapeutic Options and Molecular Targets. Vita et Sanitas, Trindade/Go 2007;1( 01).
70 - Melos JLR, Echevarria A. Sistemas Enzimáticos de Tripanossomatídeos como Potenciais Alvos. Quimioterápicos. Revista Virtual de Quimíca 2012;4(4) 374-392.
71 - Van-Griensven J, Diro E, Lopez-Velez R, Boelaert M, Lynen L, Zijlstra E, Dujardin JC, Hailu A. HIV-1 protease inhibitors for treatment of visceral leishmaniasis in HIV-co-infected individuals. Lancet Infectious Diseases 2013;13(3) 251-259. doi: 10.1016/S1473-3099(12) 70348-1.
72 - Reguera MR, Tekwani BL, Balaña-Fouce R. Poliamine transport in parasites: A potential target for new antiparasistic drug development. Comparative Biochemistry and Physiology 2005 Part C(140) 151-164.
73 - Roberts, S. C. (2013). Genetic manipulation of Leishmania parasites facilitates the exploration of the polyamine biosynthetic pathway as a potential therapeutic target. In K.V. Urbano (Ed.), Advances in Genetics Research, v.10, p. 29-54.
74 - Genestra M, Guedes SD, Souza WJS, Cysne-Finkelstein L, Soares-Bezerra RJ, Monteiro, FP, Leon LL. Nitric Oxide Synthase (NOS) Characterization in Leishmania amazonensis Axenic Amastigotes. Archives of Medical Research 2006;37(3) 328-333.
75 - Dias SS, Hogan C, Ochocka AM, Meek DW. Polo-like kinase-1 phosphorylates MDM2 at Ser260 and stimulates MDM2-mediated p53 turnover. FEBS Lett. 2009;583(22) 3543-3548. doi: 10.1016/j.febslet.2009.09.057.
76 - Lepesheva GI, Hargrove TY, Anderson S, Kleshchenko Y, Furtak V, Wawrzak Z, Villalta F, Waterman M. Sterol 14 α-demethylase as a potential target for antitrypanosomal therapy: enzyme inhibition and parasite cell growth. Chemistry and Biology 2007;14 1283–1293.
77 - Zhang K, Beverle SM. Molecular & Biochemical Parasitology Phospholipid and sphingolipid metabolism in Leishmania - Review Article. Molecular and Biochemical Parasitology 2010;170(2) 55-64.
78 - Gangjee A, Jain HD, Kurup S. Recent advances in classical and non-classical antifolates as antitumor and antiopportunistic infection agents: part I. Anticancer Agents in Medicinal Chemistry. 2007;7(5):524-42.
79 - Libusová L, Sulimenko T, Sulimenko V, Hozák P, Dráber P. Gamma- Tubulin in Leishmania: cell cycle-dependent changes in subcellular localization and heterogeneity of its isoforms, Experimental Cell Research 2004;295 (2) 375-386.
80 - Najwa, M. J, Abu-Mejdad A.; Athraa A, Al-Hilfy A. Evaluation of therapeutic effects of vicine against leishmania donovani. in vitro. Journal American Science 2013;9(5) 115-120.
81 - Singh N, Kumar M, Singh RK. Leishmaniasis: current status of available drugs and new potential drug targets. Asian Pacific Journal Tropical Medicine 2012;5(6) 485-497. doi: 10.1016/S1995-7645(12)60084-4.
82 - Takele Y, Abebe T, Weldegebreal T, Hailu A, Hailu W, Hurissa Z, Ali, J, Diro E, Sisay Y, Cloke T, Modolell M, Munder M, Tacchini-Cottier F, Müller I, Kropf P. Arginase activity in the blood of patients with visceral leishmaniasis and HIV infection. PLoS Neglected Tropical Diseases 2013;7(1):e1977. doi: 10.1371/journal.pntd.0001977.
83 - Vannier-Santos MA, Menezes D, Oliveira MF, de Mello FG The putrescine analogue 1,4-diamino-2-butanone affects polyamine synthesis, transport, ultrastructure and intracellular survival in Leishmania amazonensis. Microbiology 2008;154(Pt 10) 3104-3111.
84 - Roberts SC, Jiang Y, Gasteier J, Frydman B, Marton LJ, Heby O, Ullman B. Leishmania donovani polyamine biosynthetic enzyme overproducers as tools to investigate the mode of action of cytotoxic polyamine analogs. apy 2007;51(2) 438-445.
85 - Withers-Martinez C, Jean L, Blackman MJ. Subtilisin-like proteases of the malaria parasite. Molecular Biology 2004;53(1) 55-63.
86 - Silva-Lopez RE, Morgado-Díaz JA, Chávez MA, Giovanni-De-Simone S.Effects of serine protease inhibitors on viability and morphology of Leishmania (Leishmania) amazonensis promastigotes. Parasitolgy Research 2007;101(6) 1627-1635.
87 - Plewes KA, Barr SD, Gedamu L. Iron superoxide dismutases targeted to the glycosomes of Leishmania chagasi are important for survival. Infect Immun. 2003 Oct;71(10):5910-20.
88 - Luz, N. F.; Andrade, B. B.; Feijó, D. F.; Araújo-Santos, T.; Carvalho, G. Q.; Andrade, D.; Abánades, D. R.; Melo, E. V.; Silva, A. M.; Brodskyn, C. I.; Barral-Netto, M.; Barral, A.; Soares, R. P.; Almeida, R. P.; Bozza, M. T.; Borges, V. M. Heme oxygenase-1 promotes the persistence of Leishmania chagasi infection. The Journal of Immunology 2012;188(9):4460-7. doi: 10.4049/jimmunol.1103072.
89 - Kaur PK, Dinesh N, Soumya N, Babu NK, Singh S. Identification and characterization of a novel Ribose 5-phosphate isomerase B from Leishmania donovani. Biochemical and Biophysical Research Communications 2012;421(1) 51-56. doi: 10.1016/j.bbrc.2012.03.107.
90 - Cordeiro AT, Feliciano PR, Pinheiro MP, Nonato MC. Crystal structure of dihydroorotate dehydrogenase from Leishmania major. Biochimie 2012;94(8) 1739-48. doi: 10.1016/j.biochi.2012.04.003.
91 - Silva AM, Tavares J, Silvestre R, Ouaissi A, Coombs GH, Cordeiro-da-Silva A. Characterization of Leishmania infantum thiol-dependent reductase 1 and evaluation of its potential to induce immune protection. Parasite Immunology 2012;34(6) 345-350. doi: 10.1111/j.1365-3024.2012.01361.
92 - Rotella, D. P. (2012) Recent results in protein kinase inhibition for tropical diseases. Bioorg Med Chem Lett. Nov 15;22(22):6788-93. doi: 10.1016/j.bmcl.2012.09.044.
93 - Wetzel DM, McMahon-Pratt D, Koleske AJ. The Abl and Arg kinases mediate distinct modes of phagocytosis and are required for maximal Leishmania infection. 2012; 32(15) 3176-86. doi: 10.1128/MCB.00086-12.
94 - Cummings HE, Barbi J, Reville P, Oghumu S, Zorko N, Sarkar A, Keiser TL, Lu B, Rückle T, Varikuti S, Lezama-Davila C, Wewers MD, Whitacre C, Radzioch D, Rommel C, Seveau S, Satoskar AR. Critical role for phosphoinositide 3-kinase gamma in parasite invasion and disease progression of cutaneous leishmaniasis. Proceedings of the National Academy Sciences. U S A. 2012;109(4) 1251-6. doi: 10.1073/pnas.1110339109.
95 - Gilroy C, Olenyik T, Roberts SC, Ullman B. Spermidine synthase is required for virulence of Leishmania donovani. Infection and Immunity 2011;79(7) 2764-9, doi: 10.1128/IAI.00073-11
96 - Grover A, Katiyar SP, Singh SK, Dubey VK, Sundar D. A leishmaniasis study: structure-based screening and molecular dynamics mechanistic analysis for discovering potent inhibitors of spermidine synthase. Biochimica et Biophysica Acta 2012;1824(12):1476-83.2012doi: 10.1016/j.bbapap.2012.05.016.
97 - Mahmoudzadeh-Niknam H, McKerrow JH. Leishmania tropica: cysteine proteases are essential for growth and pathogenicity. Experimental Parasitology 2004;106 158-163.
98 - Silva-Lopez RE. Proteases de Leishmania: novos alvos para o desenvolvimento racional de fármacos. Química Nova [online] 2010;33(7) 1541-1548.
99 - Swenerton RK, Zhang S, Sajid M, Medzihradszky KF, Craik CS, Kelly BL, McKerrow JH. The oligopeptidase B of Leishmania regulates parasite enolase and immune evasion. The Journal Biology Chemistry 2011;286(1) 429-440. doi: 10.1074/jbc.M110.138313.
100 - Jäger W, Santag S, Weidner-Glunde M, Gellermann E, Kati S, Pietrek M, Viejo-Borbolla A, Schulz TF. The ubiquitin-specific protease USP7 modulates the replication of Kaposi's sarcoma-associated herpesvirus latent episomal DNA. Journal of Virology 2012;86(12) 6745-6757, doi:10.1128/JVI.06840-11.
101 - Schetelma RA, Decuypere S, T‟Kindt R, Dujardin JC, Coombs GR, Breitling G. The potential of metabolomics for Leishmania research in the post-genomics era. Parasitology 2010;137: 1291–1302.
102 - Ogungbe VI, Setzer, WN. In-silico Leishmania Target Selectivity of Antiparasitic Terpenoids. Molecules 201318(7); 7761-7847; doi:10.3390/molecules18077761.
103 - Urbina JA. Specific chemotherapy of Chagas disease: relevance, current limitations and new approaches. Acta Tropica 2010;115(1-2) 55-68.
104 - Schneider E, Hsiang YH, Liu LF. DNA topoisomerases as anticancer drug targets. Advances in Pharmacology 1990;21 149-183.
105 - Heisig P. Inhibitors of bacterial topoisomerases: mechanisms of action and resistance and clinical aspects. Planta Medica 2001;67(1) 3-12.
106 - Singh G, Jayanarayan KG, Dey CS. Novobiocin induces apoptosis-like cell death in topoisomerase II over-expressing arsenite resistant Leishmania donovani. Molecular Biochemical Parasitology 2005;141(1) 57-69.
107 - Rosypal AC, Tripp S, Lewis S, Francis J, Stoskopf MK, Larsen RS, Lindsay DS. Survey of antibodies to Trypanosoma cruzi and Leishmania spp. in gray and red fox populations from North Carolina and Virginia. Journal of Parasitology 2010;96(6) 1230-1241. doi: 10.1645/GE-2600.1.
108 - Kosec G, Alvarez VE, Agüero F, Sánchez D, Dolinar M, Turk B, Turk V, Cazzulo JJ. Metacaspases of Trypanosoma cruzi: possible candidates for programmed cell death mediators. Molecular Biochemical Parasitology 2006 Jan;145(1) 18-28.
109 - Lee N, Gannavaram S, Selvapandiyan A, Debrabant A. Characterization of metacaspases with trypsin-like activity and their putative role in programmed cell death in the protozoan parasite Leishmania. Eukaryot Cell 2007;6(10) 1745-57.
110 - Denise H, Poot J, Jiménez M, Ambit A, Herrmann DC, Vermeulen AN, Coombs GH, Mottram JC. Studies on the CPA cysteine peptidase in the Leishmania infantum genome strain JPCM5. BMC Molecular Biology 2006;13;7 42.
111 - González IJ, Desponds C, Schaff C, Mottram JC, Fasel N. Leishmania major metacaspase can replace yeast metacaspase in programmed cell death and has arginine-specific cysteine peptidase activity. International Journal of Parasitology 2007;37(2) 161-72.
112 - Meslin B, Zalila H, Fasel N, Picot S, Bienvenu AL. Are protozoan metacaspases potential parasite killers? Parasitology Vectors 2011;4:26. doi: 10.1186/1756-3305-4-26.
113 - Fritsche C, Sitz M, Wolf M, Pohl HD. Development of a defined medium for heterologous expression in Leishmania tarentolaeogy 2008;48(6) 488-95. doi: 10.1002/jobm.200700389.
114 - Iwu MM, Jackson JE, Schuster BG. Medicinal plants in the fight against leishmaniasis. Parasitology Today 1994;10 65–68.
115 - Rocha LG, Almeida JR, Maceˆdo RO, Barbosa-Filho JM. A review of natural products with antileishmanial activity. Phytomedicine 2005;12 514–535.
116 - Wang J, Peng Q, Li G. New compounds of natural resources in 2008. African Journal of Biotechnology 2009;8 4299–4307.
117 - Basso LA, Silva LHP, Fett-Neto AG, Junior WFA, Moreira IS. Palma MS, Calixto JB, Astolfi Filho S, Santos RR, Soares MBP, Santos DS. The use of biodiversity as source of new chemical entities against defined molecular targets for treatment of malaria, tuberculosis, and T-cell mediated diseases – A Review. Memórias do Instituto Oswaldo Cruz 2005;100(6) 575-606.
118 - Rates SM. Plants as source of drugs. Toxicon 2001;39 603–613.
119 - Eloff JN. Which extractant should be used for the screening and isolation of antimicrobial components from plants? Journal of Ethnopharmacology 1998;60 1–8.
120 - Beutler JA. Natural products as a foundation for drug discovery. Current Protocols in Pharmacology 2009;46(9.11) 1-9. 11.21.
121 - Sereno D, Cordeiro da Silva A, Mathieu-Daude F, Ouaissi A. Advances and perspectives in Leishmania cell based drug-screening procedures. Parasitology International 2007;56 3–7.
122 - Ajazuddin SS. Applications of novel drug delivery system for herbal formulations. Fitoterapia 2010;81(7) 680-689.
123 - Harvier P, Désormeaux A, Gagne N, Tremblay M, Poulin L, Beauchamp D, Bergeron MG. Lymphoid tissues targeting of liposome-encapsulated 2',3'-dideoxyinosine. AIDS 1995;9 701-707.
124 - Wink M. Medicinal plants: a source of anti-parasitic secondary metabolites. Molecules 2012;17(11):12771-12791.
125 - Alviano DS, Alviano CS. Plant extracts: search for new alternatives to treat microbial diseases. Current Pharmaceutical Biotechnology 2009;10(1) 106-121.
126 - Alviano DS, Barreto ALS, Dias FA, Rodrigues IA, Rosa MSS, Alviano CS, Soares RMA. Conventional therapy and promising plant-derived compounds against trypanosomatid parasites. Frontiers in Microbiology 2012;3 283. doi: 10.3389/fmicb.2012.00283. eCollection 2012.
127 - Brenzan MA, Nakamura CV, Dias Filho BP, Ueda-Nakamura T, Young MC, Coˆrrea AG, et al. Structure–activity relationship of (_) mammea A/BB derivatives against Leishmania amazonensis. Biomedical Pharmacotherapy 2008;62 651–658.
128 - Poorrajab F, Ardestani SK, Emami S, Behrouzi-Fardmoghadam M, Shafiee A, Foroumadi A. Nitroimidazolyl-1,3,4 thiadiazole-based anti-leishmanial agents: synthesis and in vitro biological evaluation. European Journal of Medicinal Chemistry 2009;44 1758–1762.
129 - Tolomeo M, Roberti M, Scapozza L, Tarantelli, C, Giacomini E, Titone L, Saporito L, Di Carlo P, Colomba C. TTAS a new stilbene derivative that induces apoptosis in Leishmania infantum. Experimental Parasitology 2013;133(1) 37-43. doi: 10.1016/j.exppara.(2012) 10.006.
130 - Volpato H, Desoti VC, Cogo J, Panice MR, Sarragiotto MH, Silva SO, Ueda-Nakamura T, Nakamura CV. The Effects of N-Butyl-1-(4-dimethylamino)phenyl-1,2,3,4-tetrahydro- β -carboline-3-carboxamide against Leishmania amazonensis Are Mediated by Mitochondrial Dysfunction. Evidence Based Complement Alternatives in Medicine 2013; 2013 874367. doi: 10.1155/2013/874367.
131 - Delfı´n DA, Morgan RE, Zhu X, Werbovetz KA. Redox-active dinitrodiphenylthioethers against Leishmania: synthesis, structure–activity relationships and mechanism of action studies. Bioorganics and Medicinal Chemistry 2009;17 820–829.
132 - Al-Qahtani A, Siddiqui YM, Bekhit AA, El-Sayed OA, Aboul-Enein HY, Al-Ahdal MN. Inhibition of growth of Leishmania donovani promastigotes by newly synthesized 1,3,4-thiadiazole analogs. Saudi Pharmacology Journal 2009;16 227–232.
133 - Srinivas N, Palne S, Nishi, Gupta S, Bhandari K. Aryloxy cyclohexyl imidazoles: a novel class of antileishmanial agents. Bioorganic Medicinal Chemical Letters 2009;15 324–327.
134 - Tonin LT, Panice MR, Nakamura CV, Rocha KJ, Santos AO, Ueda-Nakamura T, et al. Antitrypanosomal and antileishmanial activities of novel N-alkyl-(1- phenylsubstituted-beta-carboline)-3-carboxamides. Biomedical Pharmacotherapy 2010;64: 386–389.
135 - Aponte JC, Vaisberg AJ, Castillo D, Gonzalez G, Estevez Y, Arevalo J, et al. Trypanoside, anti-tuberculosis, leishmanicidal, and cytotoxic activities of tetrahydrobenzothienopyrimidines. Bioorganic Medicinal Chemistry 2010;18 2880–2886.
136 - Graebin CS, Madeira MF, Yokoyama-Yasunaka JK, Miguel DC, Uliana SR, Benitez D, et al. Synthesis and in vitro activity of limonene derivatives against Leishmania and Trypanosoma. European Journal of Medicinal Chemistry 2010;45 1524–1528.
137 - Baban B, Hansen AM, Chandler PR, Manlapat A, Bingaman A, Kahler DJ, Munn DH, Mellor AL. A minor population of splenic dendritic cells expressing CD19 mediates IDO-dependent T cell suppression via type I IFN signaling following B7 ligation. International Immunology 2005;17 909–919.
138 - Munn DH, Mellor AL. Indoleamine 2,3- dioxygenase and tumor-induced tolerance. Journal of Clinical Investigations 2007;117 1147–54.
139 - Sharma MD, Baban B, Chandler P, Hou DY, Singh N, Yagita H, Azuma M, Blazar BR, Mellor AL, Munn DH. Plasmacytoid dendritic cells from mouse tumor-draining lymph nodes directly activate mature Tregs via indoleamine 2,3-dioxygenase. Journal of Clinical Investigations 2007;117 2570–2582.
140 - Ball HJ, Yuasa HJ, Austin CJ, Weiser S, Hunt NH. Indoleamine 2,3-dioxygenase-2; a new enzyme in the kynurenine pathway. International Journal of Biochemistry & Cell Biology 2009;41(3) 467-471. doi: 10.1016/j.biocel.2008.01.005. Epub 2008 Jan 11.
141 - Metz R, Duhadaway JB, Kamasani U, Laury-Kleintop L, Muller AJ, Prendergast GC. Novel tryptophan catabolic enzyme IDO2 is the preferred biochemical target of the antitumor indoleamine 2,3-dioxygenase inhibitory compound D-1-methyl-tryptophan. Cancer Research 2007;67(15) 7082-7087.
142 - Chen W, Liang X, Peterson AJ, Munn DH, Blazar BR. The indoleamine 2,3-dioxygenase pathway is essential for human plasmacytoid dendritic cell-induced adaptive T regulatory cell generation. Journal of Immunology 2008;181(8) 5396-5404.
143 - Hou DY, Muller AJ, Sharma MD, DuHadaway J, Banerjee T, Johnson M, Mellor AL, Prendergast GC, Munn DH. Inhibition of indoleamine 2,3-dioxygenase in dendritic cells by stereoisomers of 1-methyl-tryptophan correlates with antitumor responses. Cancer Research 2007;67(2) 792-801.
144 - Johnson BA 3rd, Baban B, Mellor AL. Targeting the immunoregulatory indoleamine 2,3 dioxygenase pathway in immunotherapy. Immunotherapy. 2009;1(4) 645-661. doi: 10.2217/IMT.09.21.
145 - Cady SG, Sono M.1-Methyl-DL-tryptophan, beta-(3-benzofuranyl)-DL-alanine (the oxygen analog of tryptophan), and beta-[3-benzo(b)thienyl]-DL-alanine (the sulfur analog of tryptophan) are competitive inhibitors for indoleamine 2,3-dioxygenase. Archives of Biochemistry and Biophysics 1991;291(2) 326-333.
146 - van Griensven, J. Balasegaram M, Meheus F, Alvar J, Lynen L, Boelaert M. Combination therapy for visceral leishmaniasis. Lancet Infectious Diseases 2010;10(3) 184–194.
147 - van Griensven, J. and Diro E. Visceral leishmaniasis. Infectious Disease Clinics of North America 2012;26, 309–322.
148 - Bimal S. Sinha S, Singh SK, Narayan S, Kumar V, Verma N, Ranjan A, Sinha PK, Das VN, Pandey K, Kar SK, Das P. Leishmania donovani: CD2 biased immune response skews the SAG mediated therapy for a predominant Th1 response in experimental infection. Experimental Parasitology 2012;131 274–282.
149 - Shakya N. Sane SA, Vishwakarma P, Gupta S. Enhancement in therapeutic efficacy of miltefosine in combination with synthetic bacterial lipopeptide, Pam2Cys against experimental Visceral Leishmaniasis. Experimental Parasitology 2012;131 377–382.
150 - Sundar S. Sinha PK, Rai M, Verma DK, Nawin K, Alam S, Chakravarty J, Vaillant M, Verma N, Pandey K, Kumari P, Lal CS, Arora R, Sharma B, Ellis S, Strub-Wourgaft N, Balasegaram M, Olliaro P, Das P, Modabber F. Comparison of short-course multidrug treatment with standard therapy for visceral leishmaniasis in India: an open-label, non-inferiority, randomized controlled trial. Lancet 2011;377 477–486.
151 - Matlashewski G, Arana B, Kroeger A, Battacharya S, Sundar S, Das P, Sinha PK, Rijal S, Mondal D, Zilberstein D, Alvar J. Visceral leishmaniasis: elimination with existing interventions. Lancet Infectious Diseases 2011;11(4) 322–325.
152 - van Griensven J. and Boelaert M. Combination therapy for visceral leishmaniasis. Lancet 2011;377(9764) 443–444.