WO2015101918A1

WO2015101918A1 - Antileishmanial compositions containing fullerol and use thereof

Info

Publication number: WO2015101918A1
Application number: PCT/IB2014/067384
Authority: WO
Inventors: Frederic Jean Georges Frezard; Maurício VELOSO BRANT PINHEIRO; Maria NORMA MELO; Paula PEIXOTO CAMPOS; Marina DE SOUZA LADEIRA; Luiz Orlando Ladeira; Guilherme SANTOS RAMOS; Silvia CAROLINA GUATIMOSIN FONSECA; Kelly CRISTINA KATO; Ricardo TOSHIO FUJIWARA; Priscila GOMES DOS REIS
Original assignee: Universidade Federal De Minas Gerais - Ufmg
Priority date: 2013-12-30
Filing date: 2014-12-29
Publication date: 2015-07-09

Abstract

The present invention relates to pharmaceutical compositions containing fullerenes, preferably fullerol, which are spherical carbon nanostructures, as novel tools for the treatment of visceral and cutaneous leishmaniasis. The antileishmanial compositions containing fullerol can further comprise another leishmanicidal drug or immunomodulator, in free form or in combination with a nanocarrier, such as liposomes, which improves the pharmacokinetic proprieties of the active principles. Surprisingly, fullerol had antileishmanial activity and was able to reduce hepatotoxicity of meglumine antimoniate when administered in combination therewith in an animal model of visceral leishmaniasis. Fullerol therefore proved to be the first drug capable both of exerting antileishmanial activity without side effects and reducing the toxicity of another leishmanicidal drug without interfering in the pharmacological activity thereof. Finally, the invention comprises the use of pharmaceutical compositions containing fullerenes for the treatment of leishmaniasis.

Description

ANTILEISHMANY COMPOSITIONS CONTAINING FULEROL AND USE

The present invention relates to fullerene-containing pharmaceutical compositions, preferably fullerol, which are spherical carbon nanostructures, as novel tools for the treatment of visceral and cutaneous leishmaniasis. Fullerol-containing antileishmania compositions may comprise yet another leishmanicidal or immunomodulatory drug, either in free form or in association with a nanocarrier such as liposomes, which confers pharmacokinetic properties more favorable to the active ingredients. Fullerol has surprisingly demonstrated an antileishmania activity as well as an ability to reduce meglumine antimoniate hepatotoxicity when administered in combination with an animal model of visceral leishmaniasis. Thus, fullerol was the first drug to demonstrate the dual ability to exert antileishmania action without side effects and to reduce the toxicity of another leishmanicidal drug without interfering with its pharmacological activity. Finally, the invention comprises the use of pharmaceutical compositions containing fullerenes for the treatment of leishmaniasis.

[002] Leishmaniasis is among the most neglected tropical diseases in the world, according to the resolution of the 60th World Health Assembly in 2007. Epidemiologically, over 12 million people are infected worldwide, with 1, 3 million new people. cases reported each year, with about 20 to 30,000 deaths annually, whose illness affects the poorest populations in 88 countries (developing countries) (World Health Organization - WHO. Report of the Fifth Consultative Meeting on Leishmania / HIV Coinfection). Addis Ababa, Ethiopia, March 20 - 22, 2007; WHO Leishmaniasis, Feb. 201 3. Available at: http://www.who.int/mediacenter/factsheets/fs375/en/# Accessed: December 16, 2013).

[003] Leishmaniasis are parasitic diseases caused by more than 20 species of protozoa of the genus Leishmania spp., Of the Trypanosomatidae family, hemoflagellate parasites showing preference for phagolysosomes of the mononuclear phagocytic system. Are transmitted to humans by the bite of the vector, females of the genera Phlebotomo sp. and Lutzomyia sp. infected. The disease presents with three different clinical manifestations: visceral, or commonly known as calazar; the cutaneous; and the mucocutaneous. (RICCIARDI, LV Antileishmania Therapy: Reviewing the Past, the Present, and the Future. Gac. Med. Caracas, v.1 17, no. 2, p.93-111, 2009).

Cutaneous leishmaniasis is the most common form of leishmaniasis and presents with ulcers distributed on the exposed parts of the body, leaving lifelong scarring and severe disability. About 1/3 of the cases occur in the Americas, the Mediterranean basin, Central Asia and the Middle East. Mucocutaneous leishmaniasis leads to mutilating mucosal lesions, which consist of partial or total destruction of the mucous membranes of the nose, mouth and throat, most of them reported in Bolivia, Brazil and Peru (World Health Organization - WHO. Leishmaniasis. Feb. 2013. Available at: http: //www.who.int/mediacentre/factsheets/fs375/en/ # Accessed: December 16, 2013).

However, the most severe form of leishmaniasis comprises visceral leishmaniasis. 90% of new cases of kala-azar occur in six countries: Bangladesh, Brazil, Ethiopia, India, South Sudan and Sudan. The zoonotic form, for which dogs are the main reservoir and source of disease infection in endemic areas, is It is present in the Mediterranean Basin, China, Middle East and South America, and is caused by Leishmania infantum infantum and Leishmania infantum chagasi, the latter being the causative species of visceral leishmaniasis in Brazil (GRIENSVEN, J .; BALASEGARAM, M .; MEHEUS , F. et al., Combination therapy for visceral leishmaniasis (Lancet Infect Dis., V.10, p.184-94, 2010).

[006] The parasite's life cycle is metaxenic, where the agent undergoes transformations in the organism of the phlebotomine vector, the dipterous insect. The flagellated protozoan L. infantum is found in the mobile metacyclic promastigote form in the vector, and when phagocytized by the vertebrate host macrophages it becomes the immovable amastigote form. The cycle begins with the inoculation of infectious promastigote forms in the vertebrate host during blood meal. Upon reaching the bloodstream, promastigote forms use specific mechanisms to survive cell lysis, which will be activated by the complement system. Due to this protective mechanism, the parasite survives host attack and can still invade macrophages by manipulating cell receptors (CAMPOS-PONCE M, PONCE C, PONCE E, MAINGON RDC. Leishmania chagasi / infantum: further investigations on Leishmania tropism in atypical cutaneous and visceral leishmaniasis foci in Central America. Parasitology, New York, v. 109, p. 209-219, 2005). Within them, the parasite is protected against the host immune response, residing in the acidic environment of the secondary macrophage lysosome where it undergoes successive multiplications. Once infected, the host becomes the parasite's reservoir.

In humans, visceral leishmaniasis presents as a chronic, widespread disease characterized by irregular and long-lasting fever, hepatosplenomegaly, lymphadenopathy, anemia with leukopenia, hypergammaglobulinemia, and progressive debilitation leading to cachexia and even death. The evolution of clinical forms is diverse, ranging from spontaneous cure, oligosymptomatic and asymptomatic forms, to severe manifestations, and may reach lethality between 10% and 98% in inadequately treated and untreated cases. In recent years, the mortality of visceral leishmaniasis has been gradually increasing, from 3.6% in 1 994 to 6.7% in 2003, with an increase of 85%, and to 8.4% in 2004 (ALVARENGA, DG; ESCALDA, PMF; COSTA, ASV Visceral leishmaniasis: a retrospective study of factors associated with lethality Rev. Soe, Bras. Med. Trop., V. 43, no 2, pp. 194-97, Mar-Apr 2010 ).

Antimonial compounds were first used in the treatment of leishmaniasis in 1912 by Brazilian physician Gaspar Vianna, initially in the trivalent form. Although pentavalent antimonials such as meglumine antimoniate (Glucantime®, Sanofi-Aventis) were recognized as clinically effective in 1947, they are still the drug of choice for all leishmaniasis in most developing countries (BERMAN JD. Human leishmaniasis: clinical, diagnostic, and chemotherapeutic developments in the last 10 years (Lin. Infect. Dis. 24, 684, 1997). Amphotericin B and its various formulations are used as second-choice drugs (Ministry of Health. Visceral Leishmaniasis Surveillance and Control Manual. Brasília, 2003)

In Brazil, the therapy of leishmaniasis consists of the use of meglumine antimoniate (RICCIARDI, LV Antileishmania therapy: reviewing the past, present and future. Gac. Méd. Caracas, v.1 17, n. 2, p .93-111, 2009). However, some factors related to the pharmacokinetics of this drug disadvantage its use. Low oral absorption and high plasma clearance mean that it is administered daily parenterally (intravenously or intramuscularly), through a 20 to 40 days. In addition, side effects represent a major problem in the use of pentavalent antimonials. These appear mainly at the end of treatment and include nausea, vomiting, diarrhea, atralgias, myalgias, anorexia, elevated liver enzyme levels, electrocardiographic abnormalities, seizures, chemical pancreatitis and nephrotoxicity. Complaints from patients with pain sensation at the site and at the time of application of these medications are also common (FRÉZARD et al. Pentavalent Antimonials: New Perspectives for Old Drugs. Molecules 14, 231 7, 2009).

[010] The generally accepted model for the mechanism of action of pentavalent antimonials (SbV) is that they would be a prodrug, being reduced to the active and toxic form of trivalent antimony (Sblll), which destroys amastigote forms within the phagolysosomes. macrophages. In this context, Sblll would promote imbalance in thiol metabolism resulting in oxidative stress and apoptosis (FRÉZARD et al. Pentavalent Antimonials: New Perspectives for Old Drugs. Molecules 14, 2317, 2009).

[01 1] However, Glucantime® is contraindicated in patients taking beta-blockers and antiarrhythmic drugs with renal or hepatic impairment in pregnant women in the first two trimesters of pregnancy. There are no absolute contraindications to the use of amphotericin B; however, renal complications limit its use (Ministry of Health. Visceral Leishmaniasis Surveillance and Control Manual. Brasília, 2003).

[012] One strategy currently recommended by the World Health Organization (WHO) refers to combination treatment of different leishmanicidal drugs, with the main benefits being more effective and safer treatment and with lower risk of developing resistance (Ministry of Health Visceral Leishmaniasis Surveillance and Control Manual (Brasília, 2003). The drugs that may replace pentavalent antimonials include liposomal amphotericin B (AmBisome®), miltefosine (Impávido®) and paromomycin or aminosidine. However, the use of these drugs to date is limited, either due to the high cost and stability problem (Ambisome®), low efficacy (paromomycin), undesirable toxic effects (miltefosine, paromomycin) or the appearance of resistance (miltefosine). ) (FRÉZARD et al. Pentavalent Antimonials: New Perspectives for Old Drugs. Molecules 14, 2317, 2009). [013] Another problem in the treatment of this disease in Brazil is its high occurrence in rural areas, which makes it difficult to assist patients who have to travel to a health center to receive treatment under medical control. In this context, cases of treatment discontinuation are frequent, which may result in the emergence of drug resistance.

[014] In dogs, the main domestic reservoir of L. infantum chagasi, the use of a combination of meglumine antimoniate with allopurinol or the combination of miltefosine and allopurinol is recommended. Prolongation of treatment may be necessary and prognosis may vary from favorable to unfavorable, this will depend on the clinical condition and the immune response of the animal (DANTAS-TORRES, F.; SOLANO-GALLEGO, L; BANETH, G .; RIBEIRO, VM et al., Canine Leishmaniasis in the Old and New Worlds: Unveiled Similarities and Differences Trends in Parasitology, Oxford, v. 28, No. 12, pp. 531-538, 2012). However, regardless of the drug of choice, parasitological cure is very far from being achieved, since even if the treatment permits a reduction in the transmission of the agent through sand flies, this occurs for a short period. What will determine the effectiveness of treatment is related to the dog's immune status, pharmacokinetics and sensitivity of each Leishmania isolate, or drug resistance (AIT-OUDHIA, K .; et al. In vitro susceptibility to antimonials and amphotericin B of Leishmania infantum strains isolated from dogs in a region lacking drug selection pressure Veterinary Parasitology, Amsterdam, v 187, pp 386-393, 2012).

[015] Evidence on the ineffective pharmacological treatment of infected dogs is cumulative; They demonstrate that the animal, despite presenting clinical improvements, does not present a reversal of the infected state, which keeps the dog as a risk factor for the population and also increases the possibility of generating strains resistant to human medicines. Because there are not enough strategies to prevent transmission of the disease to humans and animals, the current most safe course of action is to sacrifice infected dogs (PANAM ERICAN HEALTH ORGANIZATION - PAHO. Surveillance, Prevention, and Control Survey of Visceral Leishmaniasis (LV)). ) in the South Cono of South America Foz do Iguazú, Brazil 2009. Available at: http://new.paho.org/hq/index.php? option = com_docman & task = doc_view & gid = 1 6961 & ltemid =, Accessed at: December 17, 2013). [016] In this context, the lack of effective treatment for canine visceral leishmaniasis represents an important limitation of current therapy. The small number of drugs available associated with the limitations found in their use points to the urgent need to develop new drugs or formulations.

[017] A successful strategy to increase the therapeutic index of drugs against visceral leishmaniasis refers to their reversible association with a carrier nanosystem, allowing them to extend their half-life in the body and direct them to the sites of infection (FRÉZARD, DEMICHELI, C. New delivery strategies for the old antimonial pentavalent drugs Expert Opinion on Drug Delivery, v. 7, pp. 1343-1358, 2010).

[018] Among the currently available drug-bearing nanosystems, liposomes occupy a prominent position. These spherical vesicles, consisting of one or more concentric lipid bilayers, may store in their internal aqueous compartment water-soluble active principles or have lipophilic or amphiphilic active principles incorporated into their membranes. Conventional liposomes are typically formed of a phospholipid, such as phosphatidylcholine, or a nonionic surfactant. They may also include in its composition cholesterol, a negatively charged phospholipid, such as phosphatidylglycerol, phosphatidic acid, diketylphosphate, and / or a positively charged phospholipid such as esterarylamine.

[019] The parenterally administered drug in liposome-encapsulated form is slowly released, thus preventing its rapid elimination by the body. The result is increased bioavailability of the drug, with potentiation of action and reduction of toxicity.

Because they are rapidly cleared from the bloodstream by macrophages of the mononuclear phagocytic system, especially the liver, spleen and bone marrow, conventional liposomes carry the drug to major sites of visceral leishmaniasis infection, which provides a greater amount of the drug to interact with the parasite. In this context, a formulation of amphotericin B in conventional liposomes (AmBisome) was developed that was successfully used to treat patients not responsive to antimonial drugs, as well as to treat patients with the PKDL form without reporting side effects. Efficacy in the 100% range in immunocompetent patients earned it US Food and Drug Administration approval (FDA) as the first liposome-based presentation to be recognized for treatment of kala azar (MEYERHOFF AUS Food and Drug Administration of AmBisome (liposomal amphotericin B) for treatment of visceral leishmaniasis. Clinical Infectious Disease, v. 28, p. 42 -48, 1999).

[021] Also in the prior art is a patent application entitled "PHARMACEUTICAL COMPOSITION UNDERSTANDING CONVENTIONAL LIPOSOMES AND PROLONGED CIRCULATION LIPOSOMES" (BR1020120052ish) for claiming the pharmaceutical composition for claiming the treatment of pharmaceutical compositions. the combination of conventional liposomes and prolonged circulation liposomes as leishmanicidal drug delivery system. This composition enables the drug to efficiently reach all sites of parasite infection, and long-cycle liposomes promote more effective targeting of leishmanicidal drugs to the bone marrow and spleen, while conventional liposomes act to target the drug to the parasite. liver.

Fullerenes were discovered in 1985 (ADAMS, PD, GROSSEKUNSTLEVE, RW, HUNG, L.-W., et al. C60: buckminsferfullerene. Nature, v. 318, p.162-163, 1985) and they represent a large family of spherical carbon nanostructures, having as a classic example the C ₆ molecule called fullerene. Due to their chemical and physical properties, these molecules have become attractive targets for use in medicine (REBECCA, M.; HSING-LIN, W .; JUN.G., et al. Impact of physicochemical properties of engineered fullerenes on key. , Toxic. And App. Pharmac, v.234, p.58-67, 2009).

[023] These nanostructures stand out for their high chemical stability and their ability to complex free radicals and promote gene expression of antioxidant enzymes, inhibiting oxidative damage (JENSEN, AW; WILSON, SR, SCHUSTER, DL Biological Applications of Fullerenes. Bioorg (Chem. V. 4, 767-79, 1996). Fullerenes have high reactivity with reactive oxygen species (ROS) such as superoxide, hydroxylated radicals and nitric oxide which attack biological macromolecules such as proteins, lipids and DNA (BOSI, S.; ROS TD; SPALLUTO, G. et al. Fullerene derivatives: an attractive tool for biological applications, v. 38, pp. 913-23, 2003). However, the technological application of fullerene in the biomedical area has been limited by the hydrophobic character of the molecule. In this In this context, a class of polyhydroxylated fullerenes, also called fullerenol or fullerol, formed by chemical modification was developed. The solubility of each fullerenol molecule is dependent on the number of hydroxyl groups introduced (KOKUBO, K. Water-Soluble Single-Nano Carbon Particles: Fullerenol and Its Derivatives. The Delivery of Nanoparticles. <Available at: http://www.intechopen .com / download / get / type / pdfs / id / 36889> Accessed December 18, 2013). Among these molecules, 22-24 hydroxyls were added to their surface, generating a hydrophilic compound called fullerol, which increases its solubility in aqueous solution and reduces its cytotoxicity (LI, J., TAKEUCHI, A., OZAWA, M., L., X., et al., Fullerol formation catalyzed by quaternary ammonium hydroxides (J. Chem. Soc., Chem. Commun, p. 1784-1785, 1993). Since then, these molecules have been studied for their therapeutic potential in different pathologies, including infectious diseases, involving oncotic oxidative stress.

[024] A water-soluble polyhydroxylated fullerene derivative, fullerenel, has shown antiviral activity in acute-infected human peripheral mononuclear cells (PBMC) and chronically HIV-1 virus-infected H9 cells with an EC50 of 7.3 ± 5, 9 μΜ and 10.8 ± 8.2 μΜ, respectively. In PBMC cells infected with HIV-2ROD, a strain resistant to antiretroviral drugs, the fullerenel had an EC50 = 0.003 ± 0.004 μΜ, thus proving to be highly effective as an antiviral drug in the treatment of this syndrome (Schinazi, RF; Sijbesma, R.; Srdanov, G.; Hill, CL; Wudl, F. Synthesis and Virucidal Activity of a Water-Soluble, Configurationally Stable, Derivatized C60 Fullerenel, Antimicrob., Agents Chemother, v. 37, pp. 1707-10. 1993).

Fullerol has been shown to act as a neuroprotective drug in oxidative stress-related degenerative diseases (DORDEVIC, A. and BOGDANOVIC, G. Fullerenol - a new nanopharmaceutic? Arch. Oncol., V.16, p.42-5 , 2008). The mechanism of fullerenol-mediated neuroprotection occurs by inhibiting glutamate receptors, and also by reducing glutamate release via increased intracellular calcium concentrations, a critical mechanism of excitotoxicity in neurons (SILVA, GA). the central nervous system, Surgical Neurology, v. 63, pp. 301-306, 2005). [026] Fullerol is not known to have acute or subacute toxicity in rodents and protects tissues from cellular damage caused by ROS following exposure to pro-oxidant toxic substances including some drugs by absorbing these free radicals that would cause tissue injury. and cellular (BOSI, S .; DA ROS, T .; SPALLUTO, G., et al. Fullerene derivatives: an attractive tool for biological applications. European Journal of Medicinal Chemistry, v. 38, p. 913-23, 2003) .

[027] A patent issued entitled "FULLERENE PHARMACEUTICAL COMPOSITIONS FOR PREVENTING OR TREATING DISORDERS" (US 6,777,445 B2) relates to a method for the prevention and treatment of bacterial infections caused by gram-positive and gram-negative bacteria. viral diseases such as dengue and viral encephalitis comprising administering pharmaceutical compositions containing a fullerene, which is administered in vivo in an amount from about 0.001 to about 100 mg / kg body weight of the subject. However, in this invention, a single water-soluble fullerene derived from the symmetrical C60 (C3) structural formula carboxylic acid, differing primarily from the present invention in that it is a fullerene with addition of 22-24 hydroxyls on the surface, is also not present. claim parenteral administration of formulations with carrier nanosystems. Moreover, it is distinguished by the use of fullerol for the treatment of visceral and cutaneous leishmaniasis.

[028] Another patent granted, US 7,947,262 B2, entitled "USE OF FULLERENES FOR THE TREATMENT OF MAST CELL AND BASOPHIL-MEDIATED DISEASE" which comprises the use of water soluble fullerene derivatives, preferably C ₄₀ -C ₈ o for the treatment of disorders involving immune system cells such as mast cells and peripheral blood basophils related to allergic and inflammatory processes such as arthritis. This invention further relates to water-soluble chimeric fullerene molecules functionalized with compounds capable of binding to receptors present on mast cells or basophils, mediating the release of cytokines and immunoglobulins, such as peptides derived from stem cell growth factors. . The patent, despite claiming liposome-encapsulated C40-C80 fullerene pharmaceutical compositions, does not preclinically and clinically evaluate, nor does it specify liposomal formulations. In addition to having a different applicability of the technology described herein. [029] The patent entitled "FINELY PARTICULATE COMPOSITE CONTAINING CARBON COMPOUND ENCAPSULATED THEREIN" (US 2007/0077432 A1), which claims compositions containing nanocomposites based on C30-C2000 closed carbon polymers, preferably between 60-120 carbon atoms, such as Fullerenes or nanotubes which may contain an associated metal, whether employed in medicine for therapy or used for diagnosis, are distinguished from the present technology in that it comprises fullerenes-containing antileishmania compositions, preferably fullerol, which are in combination with leishmanicidal or immunomodulatory drug. free or encapsulated in a colloidal carrier nanosystem.

[030] Thus, in light of what has been found in the prior art, in view of the problem of the high worldwide rate of cases of Leishmania spp. Infection, the serious side effects caused by the current meglumine antimoniate drug and the scarce number of other available drugs. For the treatment of leishmaniasis, it is concluded that the antileishmania compositions containing fullerol and other compositions containing a water soluble fullerene are promising tools to be produced and used in therapy against leishmaniasis. The present invention surprisingly reports the fullerol antileishmania activity as well as its dual ability to exert antileishmania action without side effects and to reduce the side effects of meglumine antimoniate without interfering with the pharmacological properties of meglumine when administered in combination via parenteral route. Treatment of experimentally infected mice with Leishmania infantum chagasi with free or encapsulated fullerol reduced hepatic parasitic burden in a manner equivalent to that observed by meglumine antimoniate. Fullerol showed higher antileishmania activity in the liposome encapsulated form than in the free form. However, when compared to its dosage (5 doses of 0.05 mg / kg every 4 days) with that of the first-choice medicine (20 doses of Glucantime 120 mg Sb / kg / day), fullerol was much higher. effective considering the low dose administered and the long time interval between doses. Moreover, when the treatment with liposome-encapsulated fullerol in combination with Glucantime was evaluated, the histopathological changes in the liver of the animals caused by the use of the first choice drug that characterize the hepatotoxicity were not observed. What the action demonstrates co-treatment with liposome-encapsulated fullerol. Therefore, this invention claims pharmaceutical compositions containing fullerenes as novel antileishmania drugs and their use in combination with leishmanicidal or immunomodulatory drug (s) to enhance their efficacy and reduce their toxicity.

BRIEF DESCRIPTION OF THE FIGURES

[031] Figure 1: Figure 1 comparatively shows the parasitic burden on the liver of BALB / c mice experimentally infected with Leishmania infantum (murine model of visceral leishmaniasis). The amount of parasites was determined by quantitative real-time PCR after 20 days of intraperitoneal administration with daily doses of Glucantime (120 mg Sb / kg), free fullerol (0.05 mg / kg), liposome-encapsulated fullerol. (0.05 mg / kg) and saline (negative control).

[032] Figure 2: Figure 3 shows the efficacy of treatment combining fullerol with Glucantime in a murine model of visceral leishmaniasis, with fullerol being administered in free or liposome-encapsulated form. The liver parasite burden of Leishmania infantum chagasi infected BALB / c mice, determined by quantitative real-time PCR, was evaluated after 20 days of intraperitoneal treatment with liposome-encapsulated fullerol (0.05 mg / kg / dose with 4 days interval), Glucantime (120 mg Sb / kg / day), Glucantime (120 mg Sb / kg / day) + Fullerol (0.05 mg / kg / dose / 4 days), Glucantime (120 mg Sb / kg) / day) + Liposome fullerol (0.05 mg / kg / dose / 4 days) or saline.

[033] Figure 3: Figure 3 presents the results of histopathological analysis and apoptosis by light microscopy using the HE liver staining method of Leishmania infantum experimentally infected BALB / c mice after 20 days of intraperitoneal Glucantime treatment. (120 mg Sb / kg / day), Glucantime (120 mg Sb / kg / day) associated with Fullerol (0.05 mg / kg / dose / 4 days), Glucantime (120 mg Sb / kg / day) associated with Fullerol in liposomes (0.05 mg / kg / dose / 4 days) or saline. The histopathological parameters evaluated were hydropic degeneration (A) and apoptotic index (B).

Figure 4: Figure 4 shows the viability of peritoneal macrophages exposed to fullerol. Cell viability was represented by absorbance values after 72 hours of exposure to increasing concentrations of fullerol. ^* p <0.05 = significant difference compared to the control group (0). Kruskal-Wallis test with ^Dunn's post-test.

[035] Figure 5: Figure 5 shows the in vitro antileishmania activity of fullerol and glucantime. peritoneal macrophage infection rate with L. Infantum amastigotes (BH 46 strain), 72 hours after exposure to Glucantime (0.05 mg / ml, 0.75 mg / ml or 1 mg / ml), fullerol 0.12 mg / ml . Data are shown as means ± SE. * p <0.05 = significant difference from untreated group. One-way ANOVA analysis and Bonferroni post test.

[036] Figure 6: Figure 6 shows the antileishmania activity of different doses of liposome-encapsulated fullerol in the liver of L. infantum infected BALB / c mice. In A, parasitic load in the liver of L. infantum infected BALB / c mice determined by quantitative PCR after 20 days of intraperitoneal treatment with Glucantime (120 mg Sb / kg / day), free fullerol (0.05 mg / day). kg / 4 days) and liposome-encapsulated fullerol (0.05 and 0.2 mg / kg / 4 days) and empty liposomes (same lipid dose as in the groups receiving encapsulated fullerol). The bars represent the parasite load medians (n = 6). * P <0.05 and ^*** p <0.001 Kruskal-Wallis test with Dunn 's ^post-test's. In B, proportion of mice with liver positive for Leishmania by PCR. ^** p <0.05 in Fischer's exact test.

[037] Figure 7: Figure 7 shows the antileishmania activity of different doses of liposome-encapsulated fullerol in the spleen of L. infantum infected BALB / c mice. Parasitic burden on liver of L. infantum infected BALB / c mice determined by quantitative PCR after 20 days of intraperitoneal treatment with liposome-encapsulated fullerol (0.05 and 0.2 mg / kg / 4 days), Glucantime ( 120 mg Sb / kg / day), free fullerol (0.05 mg / kg / 4 days), empty liposomes (same dose of lipid as in the groups receiving encapsulated fullerol) and saline. The bars represent the parasite load medians (n = 6). ^* P <0.05 and ^*** p <0.001, Kruskal-Wallis test with Dunn 's ^post-test's. ^* P <0.05 Mann Whitney test for comparison between Glucantime and Fui Lipo 0.2 groups.

[038] Figure 8: Figure 8 shows the liver cytokine profile of L. infantum infected BALB / c mice following treatment with different doses of liposome-encapsulated fullerol. Profile of liver cytokines determined from L. infantum infected BALB / c mice after 20 days of intraperitoneal treatment with liposome-encapsulated fullerol (0.05 and 0.2 mg / kg / 4 days), free fullerol (0.05 mg / kg / 4 days), Glucantime (120 mg Sb / kg / day), empty liposomes (same lipid dose as in the groups receiving encapsulated fullerol) and saline. Bars represent cytokine concentration averages (n = 6). ^* P <0.05, One Way ANOVA test with Bonferoni posttest.

DETAILED DESCRIPTION OF THE INVENTION

[039] The present invention relates to fullerene-containing pharmaceutical compositions as novel tools for the treatment of visceral and cutaneous leishmaniasis. Such compositions preferably comprise water-soluble fullerenes of 60 to 120 carbons with the addition of hydroxyl groups such as fullerol C ₆ oOH ₂ 2-24. Antileishmania compositions containing fullerol may comprise the drug in combination with other drug (s). leishmanicide (s) or immunomodulator (s), either in free form or in association with a nanocarrier, such as liposomes, which gives the active principles more favorable pharmacokinetic properties. Finally, the invention comprises the use of pharmaceutical compositions containing fullerenes for the treatment of leishmaniasis.

1 . Fullerol was synthesized according to the process already described in the filed patent BR 0404543-2 A2, entitled "LARGE-SCALE CONTINUOUS PRODUCTION PROCESS AND SYSTEM OF CONSTITUTED DEVICES TO PERFORM THE PROCESS". Fullerol was also encapsulated in liposomes formed of phosphatidylcholine, cholesterol and phosphatidylglycerol in a 5: 4: 1 molar ratio using the dehydration-rehydration method (FRÉZARD, F. and SCHETTINI, DA. Liposomes: physicochemical and pharmacological properties, chemotherapy applications. based on antimony Quim Nova, V. 28, No. 3, p.51 1 -518, 2005). However, these may be formed of cholesterol and a negatively or positively charged lipid in the presence or absence and polymers and / or ligands coupled to their surfaces.

2. Encapsulation was followed by calibration of vesicle size by extrusion through 200 nm pore polycarbonate membrane (NAYAR, R., HOPE, MJ, CULLIS, PR. Generation of large unilamellar vesicles from long-chain saturated phosphatidylcholines. by extrusion technique, Biochim, Biophys (Acta, v. 986, pp. 200-206, 1989). in the presence or absence of coupled polymers and / or binders on their surfaces. [040] Antileishmania activity and fullerol chemoprotective action were evaluated in Leishmania iníantum-infected BALB / c mice as a murine model of Glucantime®-treated or non-Glucantime-treated visceral leishmaniasis (meglumine antimoniate). In the first test, four groups (n = 9-10) were administered daily intraperitoneal doses for 20 days of the following treatments: 1) Glucantime at a dose of 120 mg Sb / kg body weight; 2) Fullerol at a dose of 0.05 mg / kg body weight; 3) Liposome-encapsulated fullerol at a dose of 0.05 mg / kg; 4) saline (negative control). After the treatments, the animals were sacrificed for the determination of the parasite load in the liver by the quantitative real time PCR technique.

[041] The efficacy of free and liposome-encapsulated fullerol was also evaluated after administration of only five doses (0.05 mg / kg / dose), 4 days apart (unlike the 20 doses given in the first test) compared efficacy of Glucantime (120 mg kg / day) administered daily for 20 days. The obtained result established that the use of liposome-encapsulated fullerol presented an antileishmania activity similar to that demonstrated by Glucantime, but with a more acceptable dosage.

Thus, according to the present invention, fullerol may be associated with carrier nanosystems and / or controlled release drug systems, aiming at increased stability, increased bioavailability, longer action and / or targeting to the target tissue. Such classically known carrier or controlled release systems include, but are not limited to, cyclodextrins, polymers, mucoadhesive polymers, lipid vesicles, liposomes, polymersomes, solid lipid nanoparticles, polymeric micro- and nanoparticles, micro- and nanocapsules, micro- - and nanoemulsions, dendrimers, micelles, polymer micelles, inorganic nanoparticles, carbon nanoparticles, transdermal patches, implantable polymer matrices and implantable devices.

Knowing that fullerol has antioxidant properties, the next test was to assess whether there would be any benefit from the combination of fullerol with Glucantime in terms of efficiency gains and toxicity reduction. The murine visceral leishmaniasis model was used to evaluate, by quantitative real-time PCR, the parasitic load on the liver of the animals after the treatments. A significant reduction in hepatic parasitic load was observed in the groups treated with Glucantime and Glucantime associated with liposome-encapsulated Fullerol when compared to the control group. However, the association between fullerol and Glucantime appeared to be more effective in treating leishmaniasis compared to the effect of Fullerol or Glucantime alone.

Therefore, the present invention also relates to pharmaceutical compositions comprising fullerenes associated with one or more leishmanicidal drugs and these are: pentavalent antimonials, preferably meglumine antimoniate; amphotericin B; allopurinol; ketoconazole; itraconazole; fluconazole; posaconazole; tucaresol; paramomycin; pentamidine; miltefosine, sitamaquine, iminoquimod; azithromycin; buparvaquone; tamoxifen; terbinafine; furazolidone; fluoroquinolone; domperidone; alkyl lysophospholipid or alkyl phospholipid derivative; aza-sterol; licocalcona A; maesabalid III; trichothecenes; n-acetyl cysteine; 3-substituted quinoline, or one or more immunomodulatory substances, preferably Leishmania-derived immunogenic peptides or proteins, interferon gamma, cytokines, lipid A, oligonucleotides, immunoprotectors and anti-inflammatory steroids. Livers from animals with visceral leishmaniasis submitted to antimonial chemotherapy were also used for the analysis of histopathological changes and apoptosis using the HE staining method by optical mischroscopy. Histopathological changes characteristic of hydropic degeneration were observed in the livers of animals that received Glucantime alone, showing the hepatotoxicity of the drug that can be attributed to the induction of oxidative stress. However, these changes were not observed in the group that received the combination Glucantime + liposome-encapsulated fullerol, which demonstrates the hepatoprotective action of co-treatment with fullerol in nanocarrier system.

Thus, fullerol has surprisingly demonstrated an antileishmania activity as well as an ability to reduce the hepatotoxicity caused by Glucantime when given in combination in animal model for visceral leishmaniasis. Therefore, fullerol was the first drug to demonstrate the dual ability to exert antileishmania action without side effects and to reduce the toxicity of another leishmanicidal drug without interfering with its pharmacological activity.

Thus, the set of test results allowed us to anticipate that pharmaceutical compositions containing the antimonial drug and fullerol, especially those based on liposomes, have higher antileishmania activity and reduced toxicity when compared to the commercial drug Glucantime.

Such compositions containing pure water soluble fullerenes or associated with leishmanicidal or immunomodulatory drugs may further be administered parenterally, preferably intravenously, intramuscularly, subcutaneously, intralesionally and intraperitoneally; orally, preferably in the form of capsules, dragees, tablets, syrups or elixir; topical, preferably in the form of ointment, lotion, paste or gel.

[048] In the present technology the compositions particularly comprise fullerol associated with liposome-encapsulated meglumine antimoniate.

The invention may be better understood from the following non-limiting examples.

Example 1: Fulerol antileishmania activity

[050] To assess whether Fulerol would have antileishmania activity, the parasitic load on the liver of BALB / c mice experimentally infected with Leishmania infantum (murine model of visceral leishmaniasis) was analyzed. The animals were divided into four groups (n = 9-10). Intraperitoneal doses of Glucantime (120 mg Sb / kg), free fullerol (0.05 mg / kg), liposome-encapsulated fullerol (0.05 mg / kg) and saline (negative control) were administered intraperitoneally for 20 days. . After the treatment period, the animals were sacrificed to determine the parasitic load on the liver using the quantitative real time PCR technique. Figure 1 shows a significant reduction in hepatic parasitic burden of animals treated with liposome-encapsulated Glucantime, Fulerol and Fullerol at a dose of 0.05 mg / kg ( ^* P <0.05 compared to the control group; Kruskal-Wallis test with Dunn's ^{post-test's).} Therefore, we established for the first time the anti-leishmania activity of fullerol in an experimental leishmaniasis model, which was equivalent to that of the commercial drug Glucantime.

Example 2: Effectiveness of Combining Fullerol with Glucantime in a Visceral Leishmaniasis Murine Model

[051] The liver parasite burden of Leishmania infantum-infected BALB / c mice (n = 5-6 / group) determined by quantitative real-time PCR after 20 days of intraperitoneal treatment with fullerol was evaluated. encapsulated in liposomes (0.05 mg / kg / dose 4 days apart), Glucantime (120 mg Sb / kg / day), Glucantime (120 mg Sb / kg / day) plus Fullerol (0.05 mg / kg) / dose / 4 days), Glucantime (120 mg Sb / kg / day) associated with fullerol in liposomes (0.05 mg / kg / dose / 4 days) or saline. Figure 2 shows a significant reduction in hepatic parasite burden only in the groups treated with Glucantime and Glucantime associated with liposome-encapsulated fullerol when compared to the control group (* P <0.05 with respect to the saline group; Kruskal-Wallis test with post-test ^Dunn's). However, the association between Fullerol and Glucantime appeared to be more effective in treating leishmaniasis compared to Fullerol or Glucantime alone. Therefore, this example establishes the benefit of combining Glucantime with liposome-encapsulated fullerol in terms of improved efficacy in the treatment of visceral leishmaniasis.

Example 3: Chemoprotective effect of fullerol in a visceral leishmaniasis model submitted to antimonial chemotherapy

Histopathological and apoptotic analysis was performed by light microscopy using the HE liver staining method of experimentally infected Leishmania infantum infected BALB / c mice after 20 days of intraperitoneal Glucantime treatment (120 mg Sb / kg / kg). Glucantime (120 mg Sb / kg / day) associated with fullerol (0.05 mg / kg / dose / 4 days), Glucantime (120 mg Sb / kg / day) associated with fullerol liposomes (0.05 mg / kg / dose / 4 days) or saline. The histopathological parameters evaluated were hydropic degeneration (A) and apoptotic index (B). Figure 3 shows characteristic pathological changes of hydropic degeneration and increased apoptotic index in the liver of animals receiving Glucantime showing the hepatotoxicity of the drug. These changes were also observed in the group that received the Glucantime + fullerol combination, but not in the group that received the liposome-encapsulated Glucantime + fullerol combination, demonstrating the hepatoprotective action of co-treatment with liposome-encapsulated fullerol. That is, besides exerting antileishmania action, fullerol in liposomes showed a chemoprotective action, evidenced by the reduction of Glucantime hepatotoxicity in the presence of fullerol in liposomes.

Example 4: In vitro antileishmania activity of fullerol

[053] This study was conducted in three steps. In the first stage, the cytotoxic concentration of fullerol in peritoneal macrophages was evaluated. In the second step, the antileishmania activity of fullerol in non-cytotoxic concentration for macrophages was investigated in a Leishmania infected macrophage model. In the third stage, the cytotoxic concentration of fullerol in the promatigote form of Leishmania was evaluated.

[054] For in vitro cytotoxicity and antileishmania activity tests in a Leishmania-infected macrophage model, peritoneal macrophages were isolated from BALB / c mice 3 days after intraperitoneal injection of thioglycolate 3% by peritoneal lavage with cold RPMI medium. . The cells were washed, counted and a resulting suspension of 1x10 ⁶ cells / ml density was obtained. Cells were maintained at 37 ° C in RPMI 1640 medium supplemented with 10% fetal bovine serum, streptomycin (50 pg / mL) and penicillin (1000 U / mL).

[055] In the first experiment, the cytotoxicity of fullerol in peritoneal macrophages was evaluated. For this, the cells were plated in 96-well plates at a density of 5x10 ³ cells / well and their adherence was expected to occur. The following day, cells were exposed to fullerol compounds at increasing concentrations (0 - 0.15mg / ml) and were maintained at 37 ° C in a humidified incubator and atmosphere containing 5% CO 2. After 72 hours of exposure to fullerol and glucantime, 3- [4,5-dimethylthiazol-2yl] -2,5-diphenyltetrazolium bromidetiazolyl blue (MTT-5 µg / ml) was added to each well and allowed for 4 hours. The MTT assay is based on the reduction of tetrazolium by the action of viable mitochondrial monoxygenases resulting in formazan formation (Mosman, 1983). After formation of formazan crystals, they were solubilized in DMSO and read on an Elisa reader at 570 nm. Results were shown as absorbance values and controls (not exposed to antimonial and / or fullerol).

[056] In the second experiment, cells were plated in 24-well plates containing spherical coverslips in each well at a density of 3x10 ⁵ cells / well and allowed to adhere for 1 hour. The non-adherent macrophages were then washed and 3x10 ⁶ of Leishmania infantum promastigote cells (strain MHOM / BR / 1970 / BH46) added. The infection was waited for 4 hours and treatment started by exposing the intramacrophagic amastigotes to fullerol concentration (0.12mg / ml) and increasing glucantime concentrations. As a control, untreated intramacrophagic amastigotes were used. Cells were maintained in a humidified incubator at 37 C and ^S atmosphere containing 5% C0 _2. After 72 hours of exposure At fullerol and / or glucantime, the coverslips were removed and stained in Giemsa, mounted on slides and analyzed under an optical microscope. Parasite load results were expressed as Infection Index obtained by the following formula: (% of infected macrophages) x (amastigotes / infected macrophages) / 100. Data were submitted to One Way Anova simple variance analysis followed by Bonferroni multiple comparison test.

[057] In the third experiment, Leishmania infantum promastigote cells (strain BH46) were maintained in minimal essential culture medium (a-MEM, Gibco) supplemented with 10% inactivated fetal bovine serum, 100μg / mL kanamycin and 50pg / mL ampicillin and incubated at 24 ± 1 ° C and ^at pH 7.0 in an oven BOD the promastigotes in logarithmic growth phase were placed into 96 - well plates at a density of 10 ⁶ cells / ml, exposed to increasing concentrations of 0.005 fulerol at 0.2 mg / ml. Replicates in the absence of the drug were established as controls. After 72 hours, growth was evaluated by measuring the absorbance of the cultures at 600nm using an automated microplate reading system.

[058] As shown in Figure 4, up to the concentration of 0.12 mg / ml, there is no significant effect of fullerol on cell viability. In contrast, from 0.15mg / ml the absorbance value begins to decrease significantly indicating decreased cell viability. This data allows us to infer that the maximum concentration of fullerol to be used without causing cell damage is 0.12 mg / ml.

[059] To evaluate the fullerol antileishmania activity, it was used at a maximum non-cytotoxic concentration (0.12 mg / ml) in the Leishmania infantum-infected macrophage model (strain BH46). As shown in Figure 5, fullerol alone reduced the parasite burden by about 75% compared to the control group (dark bar). The effect was equivalent to that of Glucantime given at a concentration of Sb 0.75 mg / ml.

In the study of promastigote cell growth of Leishmania infantum in the presence of increasing concentrations of fullerol in the range of 0 to 0.2 mg / ml, no cytotoxic action of fullerol was detected.

[061] In conclusion, the antileishmania action of fullerol in the Leishmania infantum-infected macrophage model has been demonstrated. The fact that fullerol did not exert cytotoxicity in the promastigote model suggests an indirect action of fullerol on the parasite through macrophage activation. Example 5: Influence of different doses of liposome-encapsulated fullerol on antileishmania activity and immune response profile in a murine model of visceral leishmaniasis

[062] This study aimed to evaluate whether administration of liposome-encapsulated fullerol at increasing doses would result in greater efficacy in a murine model of visceral leishmaniasis, as well as its possible immunomodulatory action in vivo.

In vivo efficacy evaluation of formulation treatment

[063] For efficacy testing, female BALB / c mice, aged 6-8 weeks and weighing 20-25g from the Bioterismo Center of the UFMG - CEBIO Institute of Biological Sciences, were used. The procedures used were approved by the UFMG Animal Experimentation Ethics Committee (protocol ^Q 46/2013).

Animals were infected intraperitoneally with inoculum of

2χ10 ⁷ / 100μΙ L. Infantum promastigotes (strain MHOM / BR / 1970 / BH46).

[065] Treatment began 7 days after BH46 strain inoculation and lasted 20 days. The animals were allocated to 6 experimental groups (n = 6 per group) receiving the following intraperitoneal treatments:

Group 1: treated with glucantime (120mg Sb / kg / day - 20 doses);

Group 2: treated with free fullerol in 0.15 M NaCl solution (0.05mg / kg / 4 days

- 5 doses);

Group 3: treated with 0.15 M NaCl solution containing liposome-encapsulated fullerol (0.05mg / kg / 4 days - 5 doses);

Group 4: treated with 0.15 M NaCI solution containing liposome encapsulated fullerol (0.2mg / kg / 4 days - 5 doses);

Group 5: treated with empty liposome solution (same lipid dose as in groups 4 and 5);

[071] Group 6: saline treated (control).

[072] At the end of treatment, the animals were euthanized by cervical dislocation after sedation with 8mg / kg ketamine and 5mg / kg xylazine intraperitoneally. Liver and spleen organs were collected for determination of parasite load by quantitative PCR (qPCR). [073] To determine the cytokine profile in the liver, the organs were individually weighed and homogenized in PBS containing 0.05% Tween 20 and protease inhibitors (0.1 mM phenylmethilsulfonyl fluoride; 0.1 mM benzethonium chloride; EDTA 10 mM and aprotinin A 20 Kl). After centrifugation at 8,000g for 10 min at 4 ^Q C, the supernatants were used for determination of cytokines (IL-4, IL-10, TNF-alpha and IFN-gamma) by ELISA using a commercial kit and following the manufacturer's instructions.

[074] Figure 6 (A and B) and Figure 7 show the results of parasitic burden on the liver and spleen, respectively. Significant reduction in parasite load in both liver and spleen was observed in the liposome-encapsulated fullerol (0.05 and 0.2 mg / kg) and Glucantime treated groups compared to the saline group. It is also interesting to note that free fullerol administered every 4 days did not significantly reduce parasite load and that the encapsulated form was significantly more effective than free form. Increasing the liposomal fullerol dose from 0.05 to 0.2 mg / kg resulted in a more pronounced reduction in parasitic load (Figure 6A and Figure 7), and it was not possible to detect the parasite in the liver of animals within detection of the PCR method (Figure 6B).

[075] Importantly, liposomal fullerol treatment was significantly more effective than conventional Glucantime treatment.

[076] In order to clarify the mechanism of action of fullerol in the elimination of the parasite in the liver and to show a possible immunomodulatory action, the profile of the immune response at the end of treatment was evaluated by measuring the cytokines IL-4, IL-10, TNF. alpha and IFN-gamma in the tissue. As shown in Figure 8, in the comparative analysis between treated and control (saline) groups, liposomal fullerol at a dose of 0.2 mg / kg induced a proinflammatory immune profile, with a significant increase in TNF-alpha and IFN-gamma, no significant variation of IL-10 and IL-4. On the other hand, the Glucantime group showed induction of only IL-4 and IL-10.

[077] This data shows a strong immunomodulatory action of liposomal fullerol in the liver that most likely contributed to the elimination of the parasite in this tissue. TNF-alpha is a cytokine known to stimulate hepatocytes to synthesize and secrete acute inflammatory phase proteins (mannose-linked protein, C-reactive protein) that serve to activate phagocytic complement and chemotaxis system (dendritic cells, macrophages and neutrophils). In conclusion, we have shown that: i) Liposomal fullerol administered at a dose of 0.2 mg / kg / 4 days promotes a marked reduction in parasite burden in the liver and spleen in a murine model of visceral leishmaniasis, and complete elimination of the parasite in the liver; ii) the antileishmania activity of the liposomal formulation is much higher than that of the standard drug (Glucantime); iii) the efficacy of the liposomal formulation in the liver seems to result from the induction of proinflammatory immune profile; iv) the liposomal formulation of fullerol may find applications in the treatment of other conditions affecting the liver and for which a proinflammatory immune profile would be beneficial.

Claims

1. ANTILEISHMANY PHARMACEUTICAL COMPOSITIONS characterized by comprising water soluble fullerenes.

ANTILEISHMANY PHARMACEUTICAL COMPOSITIONS according to Claim 1, characterized in that the water-soluble fullerenes have from 60 to 120 carbons with the addition of hydroxyl groups.

ANTILEISHMANE PHARMACEUTICAL COMPOSITIONS according to Claims 1 and 2, characterized in that fullerene is preferably fullerol.

(C60 OH22-24) -

ANTILEISHMANY PHARMACEUTICAL COMPOSITIONS according to Claims 1 to 3, characterized in that the water-soluble fullerenes are associated or not with one or more leishmanicidal or immunomodulatory drugs.

ANTILEISHMANIA PHARMACEUTICAL COMPOSITIONS according to claim 4, characterized in that the leishmanicidal drugs are selected from the group comprising pentavalent antimonials, preferably meglumine antimoniate; amphotericin B; allopurinol; ketoconazole; itraconazole; fluconazole; posaconazole; tucaresol; paramomycin; pentamidine; miltefosine, sitamaquine, iminoquimod; azithromycin; buparvaquone; tamoxifen; terbinafine; furazolidone; fluoroquinolone; domperidone; alkyl lysophospholipid or alkyl phospholipid derivative; aza-sterol; licocalcona A; maesabalid III; trichothecenes; n-acetyl cysteine; 3-substituted quinoline and immunomodulators are selected from the group comprising Leishmania-derived immunogenic peptides or proteins, interferon gamma, cytokines, lipid A, oligonucleotides, immunoprotectants, and steroidal anti-inflammatories.

ANTILEISHMANY PHARMACEUTICAL COMPOSITIONS according to Claim 1 to 5, characterized in that the water-soluble fullerenes are associated or not with carrier-based nanosystems and / or controlled release systems selected from the group comprising cyclodextrins, polymers, mucoadhesive polymers, lipid vesicles. , liposomes, polymersomes, solid lipid nanoparticles, polymeric micro- and nanoparticles, micro- and nanocapsules, micro- and nanoemulsions, dendrimers, micelles, polymeric micelles, inorganic nanoparticles, transdermal patches, implantable polymer matrices and implantable devices.

ANTILEISHMANY PHARMACEUTICAL COMPOSITIONS according to Claims 1 to 6, characterized in that the water-soluble fullerenes are or are not encapsulated in liposomes, and the liposomes may be composed of phospholipids, cholesterol and a negatively or positively charged lipid in the presence or absence. and of polymers and / or binders coupled to their surfaces.

ANTILEISHMANIA PHARMACEUTICAL COMPOSITIONS according to claim 7, characterized in that the liposomes are preferably composed of phosphatidylcholine, cholesterol and phosphatidylglycerol.

ANTILEISHMANIA PHARMACEUTICAL COMPOSITIONS according to claims 1 to 6, characterized in that they are administered parenterally, preferably intravenously, intramuscularly, subcutaneously, intralesionally and intraperitoneally; orally, preferably in the form of capsules, dragees, tablets, syrups or elixir; topical, preferably in the form of ointment, lotion, paste or gel.

USE OF THE ANTILEISHMANIA PHARMACEUTICAL COMPOSITIONS described in claims 1 to 9, characterized in that they are in the preparation of medicaments for the treatment of visceral, mucocutaneous and cutaneous leishmaniasis, preferably visceral leishmaniasis and the compositions preferably comprise the fullerol associated with meglumine antimoniate. encapsulated in liposomes.