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Introduction

Leishmaniasis is a type of zoonotic disease caused by more than 20 different parasite species belonging to the same genus-Leishmania. The disease is endemic majorly in tropical and subtropical regions, particularly affecting the developing countries. Leishmania is transmitted via a single vector-female Phlebotomine sand flies, to different mammalian hosts such as humans, canines, rodents etc., which are susceptible to a blood meal by the sand fly. There are clinically three forms of Leishmaniasis – 1) Visceral Leishmaniasis (Popularly known as Kala Azar) 2) Cutaneous Leishmaniasis 3) Muco-cutaneous Leishmaniasis. The disease is known to cause significant morbidity and mortality rates particularly in case of Visceral Leishmaniasis.

Available Treatments

There are majorly 5 types of drugs which are currently in use for the treatment of Leishmaniasis.

1. Pentavalent antimonial compounds- These compounds are used as the first line of treatment. The exact mechanism of action is unknown but few studies have reported the possible mechanisms such as, a) The drug has Sb (V) which is converted into Sb(III) in the presence of acidic environment of the host phagocytes. The parasite functions by maintaining the Trypanothione in its reduced state to encounter the host oxidants. The enzyme Trypanothione reductase responsible to maintain Trypanothione is inhibited by the active form Sb(III) thus leading to the impairment in the antioxidant mechanism.. b) Inhibition of type 1 topoisomerase activity. c) Reduction in ADP and GDP levels. d) Intracellular thiol level imbalance. These compounds exhibit a variety of side effects such as cardio toxicity, anorexia, nausea and hepatotoxicity. Leishmania have grown resistance against the pentavalent antimonials in India and thus alternative strategies need to be worked upon.
2. Miltefosine- The first and only oral drug available for leishmania treatment. It works by inducing apoptosis in the parasites by altering the phospholipid metabolism pertaining to the cell membrane lipids, inhibiting cytochrome C, increasing depolarization of mitochondrial membrane, production of nitric oxide in the host macrophage which is lethal to leishmania. The disadvantage with miltefosine is its teratogenecity. Clinical Cases of resistance have been reported and thus miltefosine is preferably used in combination therapies.
3. Paramomycin- Paromomycin is an antibiotic which exhibits anti leishmanial properties by targeting protein synthesis and disrupts the membrane potential of mitochondria thus leading to cell death in Leishmania. Its use majorly has been in treating the VL and CL. Paromomycin doesn’t have clinical cases of resistance but experimental resistance has been well observed making it susceptible to the former.
4. Pentamidine- The use of Pentamidine began in India after the cases of clinical resistance against antimony compounds. The mechanism of action is not clear, but it mainly inhibits mitochondrial topoisomerase II. Pentamidine possess a series of side effects such as hypoglycemia, cardiac problems, pancreatitis leading to Diabetes and gastrointestinal toxicity.
5. Amphotericin B- Amphotericin B is widely used to treat VL and is the second line of drugs recommended for the cases which have acquired resistance against the pentavalent antimony compounds. Amphotericin B shows a high affinity to bind to the major sterol of leishmania called ergosterol which is present on the membrane. The high affinity binding facilitates the formation of pores in the membrane thus disrupting the membrane organization. The membrane permeability increases leading to loss of important metabolites of the cell. Amphotericin although effective possess a list of side effects- Nephrotoxicity, chills, myocarditis etc., To counter for the side effects, a liposomal formulation of Amphotericin known as AmBisome was made commercially available which shows reduction in the side effects to a great extent and at the same time increases the bioavailability of the drug. There have been cases of resistance in clinical aspects because of the extensive use of the drug. High cost is another limiting factor in the treatment.

Challenges encountered

The current available treatment of Leishmaniasis can’t be considered gold standard due to the following challenges.

1. Development of resistant strains.
2. Alterations in the gene copy number variations of the parasite.
3. Longer duration of treatment.
4. Relapse cases and occurrence of PKDL (Post Kala Azar Dermal Leishmaniasis).
5. Painful procedures and route of administration.
6. Side effects interlinked to the treatment.
7. High cost

Thus, the need to develop alternative anti-leishmanial therapeutics arises.

Approaches of anti-leishmanial therapeutics.

1. Targeting the metabolic pathways essential for parasite survival. The exploration of the evolutionary data of Leishmania species unlocks potential targets for treatment by identifying the metabolites which exhibit structural and functional differences as compared to the host counterparts.
   • The parasite glyoxalase system- mainly responsible for detoxification, comprises of enzymes which show different substrate specificity than the enzymes present in the human glyoxalase system and thus can be targeted.
   • Protein kinases- which play a major role in cell cycle are essential for the survival and virulence of parasites. 10 MAP kinases have been reported to have distinct characteristics as compared to the host MAP kinases and thus can be exploited. One such approach has been studied to combat the antimonial resistance by upregulating MAP kinase1 gene in L donovani which leads to the increased susceptibility towards the drug.
   • Purine salvage pathway- Leishmania species lack the ability to synthesize purines from the scratch and thus they salvage the purines from the host. To do so, there are three major phosphoribosyl transferases and two transporters which transport the nucleosides into the cell. Purine reservoirs are essential for the parasitic survival and thus targeting the enzymes involved in this pathway can lead to an effective killing.
   • Antioxidant machinery of Leishmania- The major reason for survival of the amastigote form of Leishmania is its ability to combat the ROS and NO toxicity produced by the macrophages. Trypanothione reductase is one such enzyme which helps to maintain the reduced levels of trypanothione thus combating ROS. An interesting fact is although this enzyme is analogous to glutathione reductase in functionality, it is absent from the human system and thus can be considered as an effective target in production of anti-leishmanial therapeutics.
   • Cell cycle and DNA repair machinery- The important mediators of cell cycle are important for the replication and survival and discovery of inhibitors which mediate cell cycle arrest irreversibly can be a promising strategy to halt further replication. Similarly, inhibitors which inhibit the DNA repair pathways in Leishmania need to be thoroughly researched upon. A detailed understanding of the molecular mechanisms underlying the cell cycle and DNA repair machinery is an essential prerequisite for the development of novel therapeutics.
   • Topoisomerase- There are two types of topoisomerases- Type I and Type II which are further classified into Type IA, IB and IIA. These enzymes maintain the topology of the DNA. An interesting revelation of evolution led to the identification of the difference in the structure of Type IB topoisomerase. As opposed to the monomeric enzyme comprising of a single subunit in eukaryotic and prokaryotic systems like bacteria, the Type IB topoisomerase in leishmania species is comprised of heterogenic bi-subunit which becomes active when put together exhibiting DNA binding and catalytic properties. This major structural difference in the enzyme essential for survival can be a great potential target to develop anti-leishmanial therapeutics.
2. Immunotherapy- Genetic variations in individuals alter their immune response towards a variety of antigens. Particularly in case of Leishmaniasis caused by more than 20 different species, the genetic background pertaining to the species, the location and number of bites by the sand fly largely determines the immune responses. Besides, different endemic regions show different severities with respect to the same species. Thus, immune system at the host pathogen interface needs to be exploited to elevate the natural immune responses against parasites. Leishmania once inside the body, manipulates the host immune system in such a way that the two main cytokine responses oppose each other activity i.e., Th 1 responses favor the defenses against the parasite whereas Th2 responses favor the survival and proliferation of the parasites thus increasing infectivity to other parts of the body. The IFN gamma production as a result of Th1 responses induce the affected phagocytes to produce ROS and NO which can mediate oxidative killing. Thus, the role of cytokines can be exploited to create novel therapeutics that increase the levels of cytokines and chemokines responsible to curb parasitic survival.
3. Cell therapy- can be employed to make up for the tissue damage as a result of infection, the potential of mesenchymal stem cells to regenerate quickly can be exploited to account for the liver and splenic damage occurring mostly in case of visceral leishmaniasis.
4. Chemoimmunotherapy- The term chemoimmunotherapy comprises of the combination of chemotherapy with that of immune therapy wherein the former is responsible to work against the parasite and the latter is responsible to strengthen the host immunity. One such successful study has been carried out in case of the coinfection of HIV with leishmaniasis cases. The combination of liposomal amphotericin B along with the recombinant human GM-CSF improved the clinical response towards the coinfection.
5. Drug repurposing- An effective strategy to reduce the research time is to explore the clinically approved drugs for other diseases to find anti-leishmanial properties. The major advantages of drug repurposing include lesser chances of the development of resistant strains, more safety and efficacy (as it is already clinically approved), lesser side effects and cost effective in terms of research. The different bioinformatic tools and in silico-studies such as molecular docking can be used to search for the high affinity between the ligand (target molecule) and the drug. These studies can then be confirmed by various in-vivo and invitro experimentations and can be used as anti-leishmanial drugs.
6. Combination therapy- A failure in response to the monotherapy results in the cases of relapse and sometimes proves to be even fatal especially in the case of visceral leishmaniasis. Combination therapy is to use the combination of 2 drugs with one having a longer shelf life and another having the shorter shelf life. The combination therapy approach is effective in reducing the resistance against drugs and is well tolerated by the body. A study shows the combination of miltefosine and AmBisome reduced the chances of relapse and increased the immunity in a patient coinfected with HIV and VL.
7. Nanotechnology- Nanotechnology has its major application in the pipeline of drug development to increase the bioavailability of the drug and to deliver the drugs at the specific targeted site. Macrophage- the main residence of leishmania has its membrane susceptible to the entry of positively charged liposomal compounds and thus the drug encapsulated within the liposomes can be used to deliver the drug at the target site. Metal nanoparticles such as silver nanoparticles were mentioned in a study to treat cutaneous leishmaniasis.