Visceral Leishmaniasis in Latin America and therapy perspectives

Leishmaniasis visceral en América Latina y perspectivas terapéuticas

Visceral Leishmaniasis (VL) is a parasitosis caused by a flagellate protozoan of the Leishmania gender. In Latin America, the species responsible for this clinical form is Leishmania infantum; its transmission cycle involves a vector insect of the Lutzomyia genus, and reservoir animals such as dogs, which stand out for their role in domestic transmission (1).

The infectious process begins when a hematophagous female of the Lutzomyia feeds on the blood of the host releasing metacyclic promastigotes from its salivary glands, which are engulfed mainly by local macrophages, and once inside turn into amastigotes that reproduce inside the cell. After the rupture of the macrophage, the amastigotes are released and are able to infect new cells; then they migrate to the liver, bone marrow, and spleen (2) altering the spleen architecture producing self-limiting granulomatous reactions in the liver (3). Disease progress is favored by malnutrition and/or immunosuppression of the host. The disease is characterized mainly by fever, splenomegaly, hepatomegaly, lymphadenopathy, progressive cachexia, and hematological complications such as anemia, leukopenia, thrombocytopenia or pancytopenia, probably along with hypergammaglobulinemia and hypoalbuminemia, which gives rise to the need of a differential diagnosis with pathologies such as malaria, leukemia, lymphoma, and brucellosis, among others (4).

This disease has a high mortality rate in cases that do not receive treatment in a timely manner. The World Health Organization (WHO) estimates that each year, between 200,000 and 400,000 new cases of visceral leishmaniasis appear, and over 90% of them in countries such as Bangladesh, Brazil, Ethiopia, India, Sudan, and South Sudan (5). Initially, VL appeared in rural contexts, but factors such as vector domiciliation, the presence of intra and peri-domiciliary reservoirs, in addition to physical, climate, and other factors derived from human activity, have given rise to urban transmission cycles (6).

This article provides a review of this subject in the general context of the disease, emphasizing on the information found in Latin America and especially in Colombia, describes the therapeutic schemes currently approved and the perspectives in this field.

Epidemiology of Leishmaniasis in Latin America

Visceral Leishmaniasis is autochthonous in 12 American countries: the five countries with the highest number of cases in 2012 were Brazil, Paraguay, Colombia, Argentina, and Mexico, with an incidence rate of 4.8 cases per 10,000 population, and a mortality rate of 6.6% (213 deaths), showing a 1.8% reduction when compared to the previous year (7). Between 2001 and 2013, 45,490 cases were recorded, with a yearly average of 3,499 cases. The incidence of visceral leishmaniasis in the Americas for 2013 was 2.59 cases per 100,000 population, only considering the population of the area of transmission. Brazil and Paraguay were the countries with highest rates, 4.35 and 3.85 cases per 10,000 population, respectively. The death rate for that year was 6.7% (229 deaths), with reduction about one point versus the previous year (8).

With the development of the HIV-AIDS pandemic, several co-infections with other microorganisms have been observed, including Leishmania infantum. Brazil is the Latin American country with the highest co-infection rates for these two pathologies (9).

Both clinical entities potentiate each other, boosting symptom development and their complications; the most frequently observed clinical manifestations are fever and splenomegaly (10), although there have been seldom reports of atypical cutaneous lesions caused by L. infantum (L. chagasi) (11). Brazil reported 209 cases of Leishmania/HIV coinfection in 2013, accounting for 6.4% of the total cases of visceral leishmaniasis (8). Serological tests are used in the field for screening essays; however, there have been serologically negative coinfection patients, because of the immunosuppression that leads to a low production of antibodies for the parasite (12). Some authors propose the use of molecular techniques due to their high sensitivity and specificity of diagnosis of VL in this population from bone marrow aspirate or blood (13). On average, 60% of the people with VL are initially diagnosed with AIDS, in advanced immunosuppression stages, with a mean CD4+ lymphocyte count below 200/µl, which is seen in 62%-100% of the cases (14).

Epidemiología of Leishmaniasis in Colombia

According to the information of the National Health Institute (15), visceral leishmaniasis is endemic in the Valley of the Magdalena River and its affluents; some foci match vector distribution in the departments of Tolima, Huila, Cundinamarca, Bolívar, Córdoba, Sucre, Santander, and Norte de Santander.In 2015, 7,900 cases of leishmaniasis were reported, out of which 7.777 (98.4%) were cutaneous leishmaniasis, 108 cases (1.4%) were mucous leishmaniasis, and 15 cases (0.2%) were visceral leishmaniasis. When compared to 2014, there was a reduction of 3.777 cases, i.e. 32.3% variation on the epidemiologic data (16).

According to the information reported by the National System for Public Health Vigilance (SIVIGILA, for its acronym in Spanish), between 2006 and 2015, 293 cases of visceral leishmaniasis have been reported in Colombia, with an average of 29.3 cases per year. The departments involved are Bolívar, Córdoba, Huila, Risaralda, Sucre, Boyacá, Cundinamarca and Tolima, and the municipality of El Carmen de Bolívar is the one with the most cases reported (Table 1). During the 20th epidemiologic week of 2016, there have been 19 reported cases, 63% of them in the department of Bolívar.

The Colombian municipality with the largest amount of registered cases is El Carmen de Bolívar, which has contributed with 67 cases to SIVIGILA during the last 10 years, thus being the main focus of the disease, nationwide. During the last years, evidence of circulation of insects in the area has been collected, as well as vigilance on canine population (17). Recently, due to the case of a minor with symptoms compatible with VL, a focus of transmission was identified in the city of Cartagena, finding Lu. Evansi and opossums infected with L. infantum (18).

In 2012, in the municipality of Neiva, an outbreak of leishmaniasis in an urban area was reported, with a positive diagnosis for 7 minors; the reservoir study established a seropositivity of 6.1% of the analyzed canines, and complementary entomological studies confirmed the presence of Lu. Longipalpis in the peridomicile (19).

Another VL focus is located in the northern part of the department of Córdoba, in the municipalities of San Andrés de Sotavento and Tuchín, where the population is comprised of 90% indigenous persons. Children under 2 are the most affected, where malnutrition is the most significant characteristic. The only study in humans reported in this area dates back to 1995, which obtained a 40% seropositivity with Montenegro’s intradermal reaction test in children (20).

Vectors

There are about 500 known species of phlebotomines in America (21). Lu. longipalpisis the main vector of visceral leishmaniasis in Latin America, mainly located in dry tropical forests, arid and semi-arid areas, but also there are reports in very humid and forest areas (22).

In Colombia, about 164 species of Lutzomyias, 13 of them pointed out as vectors of leishmaniasis (21). Among them, Lutzomyia longipalpis and Lutzomyia evansi are the most abundant and geographically spread species. Lu. longiplapis is found in the Valley of the Magdalena River, in rural areas with tropical dry forests from the departments of Córdoba, Sucre and Bolívar, and even in suburban and urban areas predominantly in the intradomicile (23). On the other hand, Lu. evansi has been historically distributed towards the Caribbean (24). It is the main species with anthropophilic behavior in regions such as Montes de María (Sucre – Bolívar), and its adapting power has allowed it to become an urban vector for leishmaniasis (25,26). In Riohacha, department of Guajira, its circulation has been reported (27). The department of Córdoba shows evidence of transmission zones with VL cases, such as San Andrés de Sotavento (20).

However, as a result of the influence of climate change, the geographic distribution of insects may change, and the establishment of new insect niches in zones previously devoid of them this can be predicted based on environmental patterns (28). In 2013, the presence of Lu. longipalpis was reported in the department of Caldas, with an unusual altitude distribution between 392 and 1387 meters above sea level, which is evidence of the adaptation of this species due to the new climate context (29).

In the Colombian Caribbean region, there have been reports of species such as Lu. panamensis, Lu. micropyga, Lu. Shannoni and Lu. Atroclavata, with anthropophilic behavior and identification of the human food source, direct incrimination as vectors of this parasitosis requires further analysis but gives rise to the possibility of more vector species of VL involved in Colombia (25).

Reservoirs

Historically, dogs are the main reservoir of L. infantum, playing a key role in the domestic transmission and visceral form of the disease. Canines become reservoirs after a vector insect bearing the parasite bites them; some develop clinical manifestations, mainly onychogryphosis, cachexia, alopecia, and to a lesser extent, lymphadenitis, skin ulcers, and digestive disorders, while others may remain asymptomatic (30).

Within the framework of the study of disease foci, seroprevalence data has been collected in the canine population. A study in the Department of Tolima, an endemic area for this parasitosis, did not find a total correspondence between the symptoms of the dogs and the serologic studies. This could be interpreted as a limited diagnostic test for visceral leishmaniasis, and of course, the national guidelines for managing seropositive animals (30). Another interesting option for serologic tests such as screening tests, is the molecular identification of the parasite; studies in the department of Sucre report a 33.6% prevalence of Leishmania spp in canine population (31).

On the other hand, possums (Didelphis marsupialis) have been found infected with four species of Leishmania (L. infantum, L. amazonensis, L. guyanensis and L. forattinii), and are considered as a potential reservoir due to its broad distribution throughout Latin America, in both rural and urban areas (32,33,34).

Evidence provided during the last years suggest cats as an important reservoir for leishmaniasis. In Iran, a 10% infection rate with L. infantum was informed from liver and spleen samples (35); in Brazil, the natural infection with Lu. longipalpis was determined based on a cat infected with L. infantum (36). Studies in feline population must be enhanced and their role in the transmission chain of the disease must be assessed, since they are domestic animals with high presence in homes (37,38).

Pigs are another group of animals that may prove important in the anthroponotic or domestic transmission cycle of VL due to their role in the economy of rural populations; evidence of parasite specific antibodies leads to the thought that these mammals may play a role in the domestic transmission of this parasitosis. Likewise, studies conducted in endemic areas of the disease in Bangladesh report the presence of L. infantum antibodies with a 9.4% positivity in bovine cattle (37). The role of these animals as reservoirs is debated, and further structured studies are necessary in order to truly determine their role in the transmission (37).

Medicines used for treating V.L.

In a working paper published in 2013, the Panamerican Health Organization recommended the use of liposomal amphotericin B, pentavalent antimonials, and amphotericin b deoxycholate, indication periodic patient monitoring checking for warning signs of adverse effects of the medicine, as well as verifying adherence to the treatment in order to prevent resistance from appearing (39).

Currently available treatments have adverse effects that may compromise kidney, liver, and heart functions, cause hypersensitivity, and they are contraindicated under certain parameters for children under 1, people above 50, pregnancy, and immunosuppressed patients. Anti leishmanicidal therapy has an extended treatment scheme (20-30 days), and its usually intravenous route of administration leads to lack of adherence due to patient discomfort.

The currently indicated treatment in Colombia is systemic. The use of therapeutic alternatives is subject to medical criteria and specific conditions. The recommended medicines are meglumine antimoniate, miltefosine, pentamidine, and amphotericin B (Table 2). After parasitological confirmation, all patients must undergo a clinical assessment of their heart, kidney, and liver functions, and weekly follow-ups during the whole treatment (15).

Despite this situation, initiatives have been gradually promoted during the last 10 years leading to the search for new therapeutic options, some in the hands of the Drugs for Neglected Diseases initiative. This initiative involves the participation of bodies such as the Malaysian Ministry of Health; the Institute for Medical Research of Kenya; the Indian Council for Medical Research; the Oswaldo Cruz Foundation (Fiocruz) of Brazil; and the Pasteur Institut of France; Doctors without Borders; and the Special Programme for Research and Training in Tropical Diseases (TDR) of UNICEF/UNDP/World Bank, and WHO as permanent observers, and the promotion of the articulation with other public and private research entities in various countries (40).

Aiming for a combined therapy

As a result of the emerging resistance to anti leishmanicidal components, studies have been carried out in order to assess the effectiveness and cost-benefit of combined therapy. In East Africa, the effectiveness of a combined therapy of miltefosine 100 mg/day and paromomycin plus 15mg/kg/day for 10 days was assessed, achieving a >95% effectiveness; the combination of paromomycin with sodium stibogluconate for 17 days showed a 90% effectiveness (41). Cost-effectiveness analysis indicates that the miltefosine + paromomycin combination is the best cost-effective strategy when compared to other combinations including amphotericin B (42). Likewise, there are some Stege II studies to assess the safety of three therapeutic options: liposomal amphotericin B plus sodium stibogluconate, liposomal amphotericin B plus miltefosine, and only miltefosine in Ethiopia (43).

The Indian National Roadmap for Kala-Azar Elimination document published in November 2014 indicates liposomal amphotericin B in a 10 mg/kg as the first option for treating visceral leishmaniasis in that country. A second option is a combination of paromomycin miltefosine for 10 days; other options include a 28-day treatment with miltefosine or amphotericin B in multiple doses. These recommendations were approved by the WHO due to the results of a stage III study carried out between 2008 and 2010 in said country, which assessed the effectiveness of a combined therapy of liposomal amphotericin B, miltefosine, and paromomycin, and the evidence of the study achieving positive results when using a single intravenous dose of liposomal amphotericin B (40,44). The DNDi started a pharmacovigilance study in South Asia between 2012 and 2015, to assess the safety of three medicine combinations: liposomal amphotericin B + miltefosine, liposomal amphotericin B + paromomycin, and miltefosine + paromomycin. AmBisome (liposomal amphotericin B), paromomycin, and miltefosine (40); study results are not yet available, but as we mentioned above, similar studies in India and Bangladesh have yielded positive results favoring the use of these combinations.

New molecule study

Some efforts have been placed in redirecting medicines and in assessing combined therapies. Generating drugs against new therapeutic targets is a slow process that requires pre-existing as well as deeper studies of parasite cell biology, although they are always the desired objective. Ideal Target Product Profiles constitute the ideal specifications that a drug must meet considering patient needs and the main characteristics of the healthcare system involved. For the case of visceral leishmaniasis, this work route emphasizes on ideal characteristics such as the absence of adverse effects, a preference for oral administration, with a 10-day therapeutic scheme, and drug stability in tropical climates (45).

Studies developed in the last years have had the purpose of studying pharmaceutical targets such as the dihydrofolate reductase enzyme of the parasite (DHFR), cyclin-dependent kinase, ergosterol biosynthesis pathway, topoisomerase and metacaspase inhibition, and different metabolic pathways (45,46).

During the last years, the High Throughput Screening analysis technique has become more popular. This technique allows assessing large molecule collections to identify their biological activity, and a large-scale initial selection of pharmacological candidates. This methodology facilitates quantification of the ion channel activity, membrane proteins involved in several pathologies, and even promotes the development of complex clinical trials such as metabolic pathway control, monitoring intracellular events, and gene expression profiles, among others. The chemical compounds usually subject to analysis have a known pharmaceutical action. Although expensive due to the required infrastructure and equipment, these methodologies are one of the greatest perspectives in the development of new drugs (47). In the case of leishmaniasis, molecular tests have been carried out to assess their action against promastigotes in vitro, and intracellular amastigotes, generating expectations in regards to the development of metabolites with anti-kinetoplast action (48).

In Africa, sitamaquine, an 8- aminoquinoline was evaluated for treatment of the visceral form, orally administered, with promising results in stage II. However, adverse kidney effects must be assessed, as well as the development of clinical pictures such as methemoglobinemia (49).

Compounds such as chalcone, flavonoids, saponins, alkaloids, terpenoids, artemisinin, oxylipin, and guaianolides, among others, derived from plants, are included as leishmanicidal activity; some have in vitro essays and high safety indexes to develop stage I clinical trials, or to be proposed for the HTS technique. However, understanding the chemical and molecular characteristics of this type of compounds requires a large amount of time and financial investment (50).

Conclusions

Visceral leishmaniasis is a disease that mainly affects children under 7 and immunosuppressed patients; since they are vulnerable populations, Latin American government entities must sook the development of new drugs with more tolerable doses and administration routes, and less broad and severe adverse effects. Even though the path for generating new drugs demands a high investment of time and money, our continent must weigh the possibility of implementing single-dose combination therapeutic schemes that will reduce treatment duration, as has already been implemented in Asian countries after gathering the clinical evidence required for such purpose. On the other hand, deforestation, rural settlements and changes in climate parameters have favored vector insect domiciliation and its adaptation to urban environments, which would in turn allow for vector dissemination and the appearance of new disease foci; therefore, entomological vigilance is a good tool for indirect disease tracking, generating information that can predict new foci appearance.