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Title:
PHARMACEUTICAL COMPOSITIONS CONSISTING OF CYCLODEXTRIN AND AN ANTIMONY DERIVATIVE, THEIR PREPARATION AS WELL AS THEIR USE
Document Type and Number:
WIPO Patent Application WO/2006/000069
Kind Code:
A1
Abstract:
The present invention comprises processes of preparation of compounds between cyclodextrins and antimony derivatives, pharmaceutical compositions containing these compounds and methods that take advantage of these compositions for improving mainly the oral, cutaneous and percutaneous absorption of antimony. The present invention also presents therapeutic alternatives for leishmaniases and schistosomiasis based on the use of compounds between cyclodextrins and antimony derivatives by oral route and in topical application.

Inventors:
MILLAN RUBEN DARIO SINISTERRA (BR)
DEMICHELE CYNTHIA PERES (BR)
FREZARD FREDERIC JEAN GEORGES (BR)
DEMELO ALAN LANE (BR)
FERREIRA LUCAS ANTONIO MIRANDA (BR)
BEJARANO ROSEMARY OCHOA (BR)
Application Number:
PCT/BR2004/000100
Publication Date:
January 05, 2006
Filing Date:
June 24, 2004
Export Citation:
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Assignee:
UNIV MINAS GERAIS (BR)
MILLAN RUBEN DARIO SINISTERRA (BR)
DEMICHELE CYNTHIA PERES (BR)
FREZARD FREDERIC JEAN GEORGES (BR)
DEMELO ALAN LANE (BR)
FERREIRA LUCAS ANTONIO MIRANDA (BR)
BEJARANO ROSEMARY OCHOA (BR)
International Classes:
A61K47/40; A61K31/29; (IPC1-7): A61K47/40; A61K31/29
Domestic Patent References:
WO2003063788A22003-08-07
Foreign References:
JP2001233846A2001-08-28
Other References:
DEMICHELI C. ET AL: "Oral Delivery of Meglumine Antimoniate-beta-Cyclodextrin (Treatment of Leishmaniasis)", ANTIMICROB. AGENTS CHEMOTHER., vol. 48, no. 1, January 2004 (2004-01-01), pages 100 - 103
Attorney, Agent or Firm:
De Almeida, Kátia Fernandes (Almeida & Associados Rua Beneditinos 1, 11° andar -050 Rio de Janeiro - RJ, BR)
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Description:
Pharmaceutical compositions consisting of cyclodextrin and an antimony derivative, their preparation as well as their use

The present invention comprises processes of preparation of compounds between cyclodextrins and antimony derivatives, pharmaceutical compositions containing these compounds and methods that take advantage of these compositions for improving mainly the oral, cutaneous and percutaneous absorption of antimony. The present invention also presents therapeutic alternatives for leishmaniases and schistosomiasis based on the use of compounds between cyclodextrins and antimony derivatives by oral route and in topical application. Background of the Invention Antimony derivatives have been used for the treatment of leishmaniasis in humans and dogs for decades [Berman JD. 1988 Chemotherapy for leishmaniasis: biochemical mechanisms, clinical efficacy, and future strategies. Rev Infect Dis 10, 560-586] . Leishmaniases are parasitic diseases which, according to the World Health Organization (WHO), affect about 12 million people worldwide. In Brazil, recent data report the occurrence of about 20.000 new cases of the disease annually. Leishmaniases are caused by several species of flagellated protozoans belonging to the order Kinetoplastidae and to the genus Leishmania. They are zoonoses typical of tropical and subtropical rural regions of the world, although they also prevail in the suburbs of

some large cities. Leishmaniases are transmitted to

vertebrate hosts through an insect bite which releases the

parasites in the promastigote form. These parasites are

phagocytized by macrophages, inside which they transform

into amastigotes. The amastigotes multiply freely in the

acidic compartment of phagolysomes and escape the defense

systems of the host. Leishmania corresponds to a complex of

several different species that cause several types of

clinical manifestations, including cutaneous, mucocutaneous

and visceral forms. (Table 1)

Table 1. Clinical manifestations resulting from the

infection by Leishmania and world distribution of the

disease.

In Brazil, there occur tegumentar leishmaniases caused by protozoans of the Leishmania braziliensis and mexicana complexes, and visceral leishmaniasis which receives several denominations, among which are calazar, kala-azar (black fever) , tropical splenomegaly, splenomegalic anemia, and American calazar. Visceral leishmaniasis is caused by a single species of Leishmania (Leishmania) chagasi belonging to the Leishmania donovani complex. The PKDL (Post-Kala- azar dermal leishmaniasis) form is a clinical manifestation that develops in 56% of the Kala-azar patients. The dog appears as the vertebrate host of the cutaneous and visceral forms. Particularly in the case of visceral leishmaniasis, it has an important role as a reservoir and source of the disease in endemic areas. Non-clinically treated cases of visceral leishmaniasis present lethality rates of 100%. The main means to control leishmaniases in humans remains chemotherapy [Berman JD. 1997 Chemotherapy for leishmaniasis: clinical, diagnostic, and chemotherapeutic developments in the last 10 years. Clin Infect Dis 24, 684- 703] . The most used drugs are: antimony derivatives, which have been used for over 40 years in the treatment of human leishmaniases despite being potentially toxic; pentamidine and amphotericin B, which are therapeutical alternatives despite also being toxic. Among the antimony derivatives, stand out pentavalent antimonials, notably meglumine antimoniate (Glucantime, Aventis) and sodium stibogluconate (Pentostan, Welcome), which are the first-line drugs for the treatment of this disease. Due to their low oral absorption, these compounds must be administered parenterally (intravenous or intramuscular injection) for 20 to 40 days. However, the daily doses of 20 mg Sb Kg"1 need sometimes to be administered for over four months due to the increasing resistance to antimonials. This procedure has been approved for the treatment of Kala-azar and of the PKDL form in endemic areas, where resistance may occur in 5 to 70% of the patients treated with antimonials. Toxicity is another problem in the use of pentavalent antimonials. Despite their fast renal excretion, which in principle would prevent its accumulation in the organism, side effects are frequent. They usually appear at the end of the treatment and include nauseas, vomiting, diarrheas, athralgias, myalgias, anorexia, elevation of the levels of hepacellular enzymes, electrocardiographic abnormalities, convulsions, chemical pancreatitis, and nephrotoxicity [Marsden PD. 1985 Pentavalent antimonials: old drug for new diseases. Rev Soc Bras Med Trop 18, 187-198; Rodrigues MLO, Costa RS, Souza CS, Foss NT, Roselino AMF. 1999 Nephrotoxicity attributed to meglumine antimoniate (Glucantime) in the treatment of generalized cutaneous leishmaniasis. Rev Inst Med trop S. Paulo 41, 33-37] . Patients' complaints because of local pain during injection of these drugs are also common. In Brazil, the Health Ministry has reported 14 deaths caused by the use of antimonials in 2000, and this number increased to 17 in 2001 [FAPEMIG, Minas Faz Ciencias. BeIo Horizonte: FAPEMIG, no. 9, p. 12-13, Dec 2001 to Feb 2002] . In this context, WHO does not recommend a systemic treatment in the case of tegumentar leishmaniasis, except for patients with facial lesions and lesions caused by L. (V.) braziliensis to reduce the risk of mucocutaneous disease. Another limiting factor of the therapeutics of this disease in Brazil is its large occurrence in rural areas, which makes patient care difficult as they have to travel long distances to a health care center (and many times, need to remain there) to receive treatment under medical control. In this context, cases of treatment discontinuity are frequent, which tends to increase reservoirs and the incidence of drug resistance. Trivalent antimony derivatives, including tartar emetic, were the first drugs to be used in the treatment of schistosomiasis, another parasitic disease provoked by Schistosoma genus [Cioli D, Pica-Mattoccia L, Archer S. 1995 Antischistosomal drugs: past, present and future?. Pharmac Ther 68, 35-85] . Nevertheless, the use of antimony derivatives has been abandoned due to their undesirable toxic effects and to the appearance of the orally-active and less toxic drugs, oxamniquine and praziquentel. At present, the production of oxamniquine has been interrupted and the appearance of parasites resistant to praziquentel is seen as an emerging problem. In face of these limitations, the WHO, along with other related institutions such as TDR, recommends and supports the search of new drugs and formulations as well as simpler and safer administration routes such as the oral and topical routes, both for the treatment of leishmaniasis [Leishmaniasis. In: UNDP/World Bank/WHO Special Programme For Research And Training In Tropical Diseases (TDR) . Tropical Disease Research Programme Report, 11; progress 1991-92. Geneva: World Health Organization, 1993. p. 77] and the treatment of schistosomiasis. Among the most promising approaches that have been presented so far in the state of the art for the treatment of leishmaniases, one can mention those based on the use of liposomes, of lipophilic substances active by oral route and/or in topical application, of formulations for local therapy, and of synergetic combinations of drugs. The use of liposomes as drug carriers is a new trend in the pharmaceutical industry and allowed the development of novel leishmanicidal drugs. These spherical vesicles made up of one or several concentric lipid bilayers can store hydrosoluble active compounds in their inner aqueous compartment, or have lipophilic or amphiphilic active compounds incorporated in their membranes. As a result, the drug is released slowly, thus avoiding its rapid elimination by the organism. The main benefits are the increased drug bioavailability, the potentialization of its action and the reduction of its toxicity. In the case of visceral leishmaniasis, as the vesicles are quickly captured by macrophages, they naturally target the drug to the infection sites, which makes a larger quantity of the drug available to interact with the parasite. In this context, a preparation of amphotericin B encapsulated in liposomes (AmBisome®) [WO9640060-A1] was developed and used successfully in the treatment of patients non- responsive to antimonial drugs as well as of patients with the PKDL form without any reported side effects. Its efficacy in the range of 100% with immunocompetent patients led to its approval by the U.S. Food and Drug Administration (FDA) as the first liposome-based formulation for the treatment of Kala-azar [Meyerhoff A. 1999 U.S. Food and Drug Administration approval of AmBisome (liposomal amphotericin B) for treatment of visceral leishmaniasis. Clin Infect Dis 28, 42-48] . In the case of antimonials, liposomal compositions have also been developed [US4186183A; EP72234A; WO9604890-A1; US4594241] . In experimental models of Kala-azar, these preparations were found to be at least 200 times more effective than non-encapsulated antimonial. Nevertheless, the low stability of these formulations and reports of toxicity have limited the development of these formulations [Frezard F, Michalick MS, Soares CF, Demicheli C. 2000 Novel methods for the encapsulation of meglumine antimoniate into liposomes. Braz J Med Biol Res 33, 841-846] . Another limiting factor for the use of liposome-based formulations in poor countries is their high cost. Active compounds with lipophilic character may also be administered orally and/or applied topically. This is the case of phospholipids containing sulfur in their composition [WO9402153-A1] , of phosphoethanolamine derivatives [EP534445-A1] and of phosphocholine derivatives [WO90937289-A1; EP507337-A2; DE19835611-A1] . One of the phosphocholine derivatives, hexadecylphosphocholine (or miltefosine) , previously evaluated for cancer treatment, is now being clinically tested in India for the oral treatment of Kala-azar [Sundar S, Jha TK, Thakur CP, Engel J, Sendermann H, Fischer C, Junke K, Bryceson A, Berman J. 2002 Oral miltefosine for Indian Visceral leishmaniasis. N Engl J Med 347, 1739-1746]. Preliminary results of these studies show a high efficacy of this drug by oral route; however, undesirable side-effects have also been reported. Among the topical formulations tested, the preparation containing paromomycin (or aminosidine) is one of the most effective in the treatment of cutaneous leishmaniasis [Gamier T, Croft SL. 2002 Topical treatment for cutaneous leishmaniasis. Curr Opin Investig Drugs 3, 538-544] . It is noteworthy that topical formulations of the antimonial drug, pentostan, did not present satisfactory therapeutical response [Costa JM, Barrios LA, Netto EM, Marsden PD. 1986 Topical pentostam in an attempt to produce more rapid healing of skin ulcers due to Leishmania braziliensis braziliensis. Rev Soc Bras Med Trop 19, 199-200] . Nevertheless, the efficacy of intralesional meglumine antimoniate [Alkhawajah AM, Larbi E, al-Gindan Y, Abahussein A, Jain S. 1997 Treatment of cutaneous leishmaniasis with antimony: intramuscular versus intralesional administration. Ann Trop Med Parasitol 91, 899-905] suggests that local therapy, based on antimonials, is a promising approach for the treatment of tegumentar leishmaniasis. Several leishmanicidal drug combinations have shown to be synergetic. The association of gentamycin to paromomycin increased the efficacy of paromomycin in topical application [WO9406439-A1] . In turn, the combination of aminosidine with sodium stibogluconate was found to be an effective means of treating the visceral forms non- responsive to conventional treatment. Similarly, the association of this antimonial with another drug under clinical evaluation, allopurinol, has shown to be effective even in cases of antimonial resistance [Martinez S, Gonzalez M, Vernaza ME. 1997 Treatment of cutaneous leishmaniasis with allopurinol and stibogluconate: Clin Infect Dis 24, 165-169; Leishmaniasis. In: UNDP/World Bank/WHO Special Programme For Research And Training In Tropical Diseases (TDR) . Tropical Disease Research Programme Report, 13; progress 1995-96. Geneva: World Health Organization, 1997. cap. 8, p. 100-111] . Immunochemotherapy, which associates immunomodulators to pentavalent antimonials, was found to be an effective means of reducing the antimonial dose applied, while maintaining the treatment efficacy [Murray HW, Berman JD, Wright SD. 1988 Immunochemotherapy for intracellular Leishmania donovani infection: gamma interferon plus pentavalent antimony. J Infect Dis 157, 973-978; Machado-Pinto J, Pinto J, da Costa CA, Genaro O, Marques MJ, Modabber F, Mayrink W. 2002 Immunochemotherapy for cutaneous leishmaniasis: a controlled trial using killed Leishmania (Leishmania) amazonensis vaccine plus antimonial. Int J Dermatol 41, 73- 78] . In summary, the state of the art shows that antimonial therapy suffers several limitations, the most serious ones being the need of parenteral administration, its undesirable toxic effects, and the development of resistance. Conversely, therapeutical alternatives have been explored, however, the development of these new products has been limited so far by their high cost, their low efficacy or their undesirable side effects. It is also known in the present state of the art that a drug may undergo changes of properties such as solubility, biodistribution, and pharmacokinetics due to its interaction with appropriate encapsulating agents such as cyclodextrins and liposomes. Cyclodextrins are cyclical oligosaccharides that include six, seven, or eight glucopyranose units called alfa (α) , beta (β) and gamma (γ) cyclodextrin, respectively. Due to steric interactions, cyclodextrins form a structure shaped like a truncated cone with a polar surface and an internal apolar cavity. The external surface of cyclodextrins is constituted by hydroxyls bonded to C-2, C-3, and C-6, which allow solvation by water molecules as well as the introduction of substituents without altering the internal cavity. Primary hydroxyls are located in the narrower side of the cone, while secondary hydroxyls are located in the larger side. Despite the stability conferred to the cone by the intramolecular hydrogen bonds, it is flexible enough to allow substantial modifications from its regular shape. β-cyclodextrin is most used due to considerations of cost, availability, approval by health authorities, and cavity dimension. Nevertheless, this cyclodextrin presents low water solubility as a result of the formation of intramolecular hydrogen bonds between the hydroxyls of C-2 and C-3. This difficulty can be circumvented with the preparation of hydrophilic derivatives, such as hydroxyalkylated cyclodextrins. Cyclodextrins can form complexes both with apolar substances, which will be partially or completely included in its apolar cavity, and with polar substances, which will interact with the surface of cyclodextrin [WO200232459-A2] . In this way, cyclodextrins (hosts) allow the modulation of solubility of substances (guests) in aqueous solutions, usually by increasing it for apolar molecules and reducing it for polar molecules. The state of the art reports an increase in the bioavailability of drugs by oral, nasal, rectal, and topical routes through their inclusion in the apolar cavity of cyclodextrins [Szejtli J. 1998 Introduction and general overview of cyclodextrin chemistry. Chemical Reviews 98, 1743-1754; Szejtli J. 1997 Utilization of cyclodextrins in industrial products and processes. J Mater Chem 7, 575-587; Loftsson T, Masson M. 2001 Cyclodextrin in topical drug formulations: theory and practice. Int J Pharm 225, 15-30] . According to detailed toxicity, mutagenesis, teratogenesis, and carcinogenesis studies on cyclodextrins [Rajewski RA, Stella V. 1996 Pharmaceutical applications of cyclodextrins. 2. In vivo drug delivery. J Pharm Sci 85, 1142-1169] , they present low toxicity, particularly hydroxypropyl-β-cyclodextrin [Szejtli J. 1990 Cyclodextrins: Properties and applications. Drug Investig 2 (S.4), 11-21] . The use of cyclodextrins as food additives has already been authorized in countries like Japan and Hungary, and for more specific applications in France and Denmark. In addition, they are obtained from a renewable source through the degradation of starch. All these characteristics turn this system greatly attractive for the investigation of new applications. The state of the art reports inventions [JP2000264903; JP2001233846; JP2001213836] characterized by the preparation of composites, which comprises cyclodextrin and metallic salts of perfluoroalkyl sulfonic acid, of bis- perfluoroalkyl sulfonyl imide or of tris-perfluoroalkyl sulfonyl methyl, to be used as Lewis acid catalysts. These composites include antimony derivatives, but are limited to some perfluoroalkylated derivatives. In addition, applications of these compounds have been proposed in the field of catalysis but not in the pharmaceutical field, which characterizes the present invention. One also encounters, in the state of the art, the preparation of polyglycolic-based biomaterials from purified and dry glycolide and cyclodextrin, using antimony trioxide as a catalyst [KR9592607] . However, antimony does not enter in the final composition of these biomaterials, contrary to the preparations of the present invention. The state of art also reports the preparation processes of liposomes, which encapsulate cyclodextrin- based association or inclusion compounds [WO9515746; WO9423697; WO9704747; McCormack B, Gregoriadis G. 1994 Entrapment of cyclodextrin-drug complexes into Liposomes: Potential advantages in drug delivery. J Drug Targeting 2, 449-455] . Considering that liposomes [WO8701938; EP224837; Egbaria K, Weiner N. 1990 Liposomes as a topical drug delivery system. Adv Drug Deliv Rev 5, 287-300] and cyclodextrins both promote cutaneous and transdermal drug absorption, it can be expected a large efficacy from the association of these two systems in the control of percutaneous absorption. The present invention comprises processes of preparation of compounds between cyclodextrins or their derivatives and antimony or its derivatives as well as pharmaceutical compositions containing these compounds. These processes include the reaction of cyclodextrins with antimony oxides, obtained either from potassium hexahydroxoantimonate (KSb(OH)6) or other alkaline, alkalino-terrous or rare hearth metals, or from the hydrolysis of antimony pentachloride (SbCl5) in water. These processes also include the mixture of antimony derivatives with cyclodextrins in aqueous solution or in solid state. Another characteristic of the present invention is the preparation of different antimony derivatives in association with different cyclodextrins or derivatives. Antimony derivatives include preferentially, but are not limited to, the pentavalent antimonials currently used in the treatment of leishmaniasis, meglumine antimoniate and sodium stibogluconate, but also in a broader way, antimony complexes obtained from any type of sugars, poly-sugars, aliphatic alcohols, acids and amines derived from sugars and alcohols or from complexing agents that contain sulfhydryl, hydroxy!, carboxylate, and sulfate groups or from nucleosides or nucleoside derivatives, as for example riboside allopurinol, and/or mixtures thereof. Cyclodextrins include α~, β-, γ-cyclodextrins. Cyclodextrin derivatives can be selected from the group containing alkyl, hydroxyalkyl, hydroxypropyl, and acyl, or cross- linked cyclodextrins or cyclodextrin polymers and/or their mixtures in aqueous solution or in solid state. The present invention also comprises the preparation of pharmaceutical compositions from these compounds based on the association of antimony and cyclodextrins, including liposomes compositions encapsulating the resulting compounds. In agreement with the present invention, it has been demonstrated that the association of an antimonial to cyclodextrin results in increased oral, cutaneous and percutaneous absorption of antimony. This increase in antimony bioavailability characterizes the compounds and compositions of the present invention as potential drugs for use by oral route and in topical application in the treatment of leishmaniases and schistosomiasis. It is important to point out that one of the compounds of the present invention, meglumine antimoniate associated to β-cyclodextrin, was found to be active by oral route, when administered to mice infected by Leishmania amazonensis, an experimental model of cutaneous leishmaniasis [Demicheli C, Ochoa R, Bento JBB, Falcao CA, Rossi-Bergmann B, Sinisterra RD, Frezard F. 2004 Oral delivery of meglumine antimoniate-β-cyclodextrin for treatment of leishmaniasis. Antimicrob Agents Chemother 48, 100-103] . The leishmanicidal activity of this compound by oral route was equivalent to that achieved with the administration of the antimonial in its free form given by parenteral (intraperitoneal) route at a twofold higher dose of antimony. Conversely, the activity of the association compound was significantly higher than that of the antimonial in the free form given by oral route at a fourfold higher antimony dose. Therefore, this invention reports for the first time an antimony-based composition that shows effectiveness by oral route against leishmaniasis. It is noteworthy that it has not been found, in the state of the art, any invention that reports the preparation of compounds between cyclodextrins or their derivatives and antimony or its derivatives, as well as pharmaceutical compositions, aiming the treatment of leishmaniases and schistosomiasis. The leishmanicidal compositions of the present invention, contrary to those reported so far in the state of the art, are not related to the development of a new drug, but rather to the renovation an old drug (the antimonial) already in clinical use for decades, which represents a major advantage in terms of applicability. A second advantage, with respect to other therapeutic alternatives proposed in the state of the art, is that antimonials have recognized and elevated effectiveness against all forms of leishmaniasis. A third advantage of the present invention is the use of the oral and topical routes of administration. It is worth pointing out that oral and topical pharmaceutical compositions based on the association of an antimonial to cyclodextrin, as proposed in the present invention, represent economically-advantageous products for the pharmaceutical industries, considering that the cost of preparation of oral or topical formulations is lower than those of injectable formulations. As other benefits, the present invention should result in a simpler and potentially less toxic treatment of leishmaniasis and schistosomiasis, in the reduction of public expenses with the treatment of leishmaniasis, in the increase in patient compliance and in the reduction of risk of increase in the number of cases of resistance. The present invention is also related to the use of these new compounds or formulations in association with other pharmacologically active compounds such as substances with immunomodulating activity. The present invention can be better understood through the following non-limiting examples. Example 1 Preparation of a compound between β-cyclodextrin and meglumine antimoniate. Initially, it was added 0.001 mol of meglumine antimoniate to 40 mL of water containing 0.001 mol of β- cyclodextrin (β-CD) dissolved. The mixture was kept under stirring at a temperature ranging from 40 to 70°C for a period from 48 to 96 hours. Next, the solvent was eliminated from the resulting solution, by freeze-drying for example. A meglumine antimoniate/β-cyclodextrin (MA/β- CD) association compound, showing a (1:1) stoichiometry and a molecular formula of was then obtained (98% yield) . The product was characterized through elemental analysis (C, H, N), atomic absorption (Sb), thermal analysis, and HPLC (High Performance Liquid Chromatography) . Elemental analysis yielded: %C = 33.51 (33.89); %H = 6.57 (6.57); %N = 0.78 (0.81); %Sb = 7.94 (7.02) . Thermogravimetry indicated the presence of nine H2O molecules. The reaction product was evaluated by HPLC using an HRC-NH2 column (5 mm; 250 x 4.6 mm; Shimadzu) as stationary phase, and a mixture of acetonitrile/water (1:1, v/v) flux= lmL/min as mobile phase and a refraction index detector (Waters-410) . The retention time obtained for the resulting product was compared to those of meglumine antimoniate (MA) , β-cyclodextrin (β-CD) , and to the mixture of MA and β-CD at 1:1 molar ratio. The results are shown in Table 2. These data indicate that the MA/β-CD association compound does not present the retention times characteristic of MA, while the mixture made at the same molar ratio presents retention times characteristic of MA peaks and of β-CD peak. These data clearly indicate that the synthesized compound is not a mixture in solution, but rather an association compound of MA and β-CD. Table 2. HPLC retention times of meglumine antimoniate (MA) , β-cyclodextrin (β-CD) , the association compound MA/β- CD, and the equimolar mixture of the two components (MA + β-CD) . Species Retention time (min) MA/β-CD 4.49 β-CD 4.58 MA 6.43 6.62 MA + β-CD 4.57 6.42

The mode of interaction of MA with β-cyclodextrin in the MA/β-CD compound was investigated by proton nuclear magnetic resonance (1H NMR) . The chemical shifts of β- cyclodextrin protons were compared between MA/β-CD and free β-cyclodextrin (in D2O and DMSO-dβ) • The most pronounced differences were observed for the hydrogens lying outside the cyclodextrin cavity, suggesting that MA interacts with hydroxyls located on the external face of cyclodextrin. This result also indicates that MA/β-CD compound is not an inclusion compound, but rather an association compound. Example 2 Preparation of a compound between D-cyclodextrin and pentavalent antimony from SbCIs- To 25.0 mL of distilled water at room temperature, 0.200 g of β-CD was added. An equimolar quantity of hydrated antimony pentoxide obtained from the hydrolysis of SbCl5 in water was then added to this solution. The system was stirred and heated to 6O0C for 3 hours. The pH of the solution was kept around 7 with potassium hydroxide. After heating, the system was centrifuged to eliminate the unreacted antimony oxide. The complex was precipitated with acetone. An antimony complex, showing a stoichiometry of 1:1 and molecular formula of KC42H84O46Sb, was then obtained (yield of 95%) . The product was characterized by elemental analyses (C, H, N) and atomic absorption (Sb) . Elemental analyses yielded: %C =32.83 (33.90); %H = 5.70 (5.70); %Sb =8.27 (8.19). Example 3 Preparation of a compound between D-cyclodextrin and pentavalent antimony from KSb(OH)6. To 25.0 mL of distilled water at room temperature, 0.200 g of β-CD was added. To this solution, KSb(OH)6 was added at Sb/β-CD molar ratio of 1:1. The system was stirred and heated to 600C for 3 hours. The pH of the solution was kept around 7 with potassium hydroxide. The complex was precipitated with acetone. An antimony complex, showing a stoichiometry of 1:1 and a molecular formula of KC42H84O46Sb was then obtained (yield of 96%) . The product was characterized by elemental analyses (C, H, N) , and atomic absorption (Sb) . Elemental analyses yielded: %C =32.00 (33.90); %H = 5.61 (5.70); %Sb =8.01 (8.19). Example 4 Preparation and characterization of liposomes containing the MA/β-CD association compound. Multilamellar vesicles (MLVs) were prepared using 25 mg of distearoylphosphatidylcholine (DSPC) in 2 mL deionized water. Next, the MLVs suspension was submitted to ultrasonification at 55°C. The resulting suspension of small unilamellar vesicles was mixed with 8 mL aqueous solution of meglumine antimoniate or of MA/β-CD compound, in both cases containing 1.5 mg Sb. These suspensions were immediately frozen and subsequently dried for 24 hours. The dry powder was rehydrated according to the following protocol. 0.2 mL of deionized water was added and the mixture was incubated for 30 minutes at 6O0C. Next, an equal volume of phosphate buffer saline (PBS: 150 mM NaCl, 10 mM phosphate, pH 7.4) was added and the mixture was incubated for 30 min at 6O0C. Finally, 1 mL of PBS was added and the mixture was incubated for 30 min at 600C. Liposomes were separated from non-encapsulated compound by centrifugation (10,000Xg, 25°C, 20 min) . The liposome pellet was then washed twice with PBS and re-suspended in 2 mL of the same solution. Antimony concentration was determined in the resulting liposome suspension by plasma emission spectroscopy, after digestion of the sample with nitric acid. The encapsulation efficiency of antimony and the encapsulated antimony-to-lipid ratio are presented in Table 3. Table 3 - Characteristics of encapsulation of antimony in liposome from meglumine antimoniate (MA) and its association compound (MA/β-CD) .

Compound Encapsulation Sb/lipid final ratio efficiency (w/w) MA 46 % 0.027 MA/β-CD 70 % 0.042

Significant increases in both the encapsulation efficiency and Sb/lipid ratio were observed, when MA was associated to β-CD. It can be concluded that the MA/β-CD association compound can be encapsulated in liposomes, and that the association of MA to β-CD results in an increase in the encapsulation efficiency of antimony. Example 5 In vitro percutaneous absorption of antimony in vitro from meglumine antimoniate and its association compound with β-cyclodextrin. The experiment was carried out in Franz diffusion cells made up of an upper and a lower compartment with volume of 6.7 cm3 and membrane surface area of 1.76 cm2. Hairless male mouse skins (HRS/J strain with 60-70 days) were used. Experiments were carried out in triplicate in the presence and in the absence of stratum corneum (SC) . The skin with stratum corneum represents an intact skin, while that without stratum corneum represents a hypertrophic skin, a situation commonly observed in the histopathological evolution of cutaneous leishmaniasis. The receptor compartment was filled with phosphate buffer saline at pH 7.4. The receptor fluid was kept at 37 ± 0.5 0C and stirred continuously with a magnetic bar. 0.1 mL of the preparations (either MA. or MA/β -CD complex, both containing 430 μg of antimony) was applied onto the skin. The donor compartment was kept open to allow the evaporation of the volatile aqueous phase of the preparations, simulating normal conditions of use. The samples were collected at time intervals of 2, 4, 6, and 8 hours, through the total removal of the receptor fluid and filling the receptor compartment with a new solution. To determine the quantity of antimony absorbed by skin, the preparation was removed from the skin surface using 0.5 mL of aqueous solution at 1% non-ionic surfactant (polyoxyethylene 20 oleyl ether) followed by application of distilled water. This procedure was repeated twice. The skin surface was dried with the help of a q-tip and kept in 5 mL phosphate buffer pH 8.0 for 12 hours. After this period, the tissue was dilacerated (UltraTurrax operating at 12.000 rpm for 5 rain) and the samples were centrifuged for tissue removal. Antimony concentration was determined in the receptor fluid and in the skin extract by electrothermal atomic absorption spectrometry with graphite furnace (ETAAS), using a ruthenium and rhodium mixture as permanent modifier. The results obtained are presented in Table 4. Table 4 - Cutaneous and percutaneous absorption of antimony across mouse skin with and without stratum corneum from meglumine antimoniate (MA) and its association compound with β-cyclodextrin (MA/β-CD) . The values are given as average of the permeated quantity ± standard deviation (n =

Quantity of permeated antimony (Dg) Time With stratum corneum Without stratum corneum (hours) MZV MA/β-CD MA MA/β-CD 2 7.7 ± 4.1 22.0 ± 5.0 15 ± 0.5 125 ± 6 4 12.3 + 2.9 29.3 ± 8.1 27 + 1.5 166 ± 2 6 22.0 ± 6.3 48.3 ± 11.9 45 ± 2.0 187 ± 1 8 30.0 ± 8.0 61.7 ± 11.7 80 ± 4.0 217 + 1

When compared to MA, the MA/β-CD association compound, object of the present invention, showed an increased percutaneous absorption of antimony, both in the presence and absence of stratum corneum. The quantities of antimony- transported across the intact skin after 8 hours represent 7% and 14% of the antimony dose initially applied in the case of MA and MA/β-CD, respectively. Therefore, the association of MA with β-CD resulted in an antimony flux approximately twofold larger across the skin. It is worth pointing out that the quantity of antimony absorbed by the skin also increased significantly. The quantities of antimony found in skin 8 hours after the application of MA and MA/β-CD were 20.3 ± 1.5 μg and 27 ± 2 μg Sb, respectively. Antimony permeation was larger in skin without stratum corneum than in intact skin for both compounds. Nevertheless, the association of MA with β-CD resulted in an antimony flow approximately sixfold to eightfold larger when compared to that of free MA. It can be concluded that the association of the antimony derivative to β-CD resulted in increased cutaneous and percutaneous absorption of antimony. This increase in bioavailability of antimony shows the great potential of these preparations for the topical treatment of leishmaniasis. Example 6 Oral absorption of antimony in mice from meglumine antimoniate and its association compound with D- cyclodextrin. Female SWISS mice weighing 30 ± 3 g received a dose of 82.5 mg Sb/kg body weight of meglumine antimoniate or its association compound with D-cyclodextrin in aqueous solution by oral route. At different times after administration (0.5, 1, 2, 3, 4, 6, 12, 24 hours), mice were sacrificed (3 animals in each group and at each time) . Blood samples were collected and sera were separated. Antimony concentration in serum was determined by ETAAS with graphite furnace. The results obtained are presented in Table 5. Table 5. Antimony concentrations in mouse serum (μg/L) at different times after oral administration of a single dose (83 mg Sb/kg body weight) of meglumine antimoniate (MA) or its association compound with β-cyclodextrin (MA/β-CD) . The values are given as average ± standard deviation (n = 3 animals at each time) . Time (hours) MA MA/β-CD 0.5 699 ± 72 3781 ± 896 1 613 ± 61 1972 ± 411 2 396 ± 11 1346 ± 92 3 356 ± 8 691 ± 35 4 109 ± 8 677 ± 58 6 58 + 18 244 ± 65 12 30 ± 11 101 ± 21 24 na* Na* * na, below detection limit. When compared to MA, the MA/β-CD association compound, object of the present invention, showed an increased oral absorption of antimony. It can be concluded that the association of the antimony derivative to β-CD resulted in enhanced bioavailability of antimony by oral route, showing the great potential of these compounds in the treatment of leishmaniasis by this administration route. Example 7 Oral absorption of antimony in mice from meglumine antimoniate and its association compound with 2- hydroxypropyl-β-cyclodextrin. Female SWISS mice weighing 30 ± 3 g received by oral route a single dose of 82.5 mg Sb/kg body weight of meglumine antimoniate or its association compound with 2- hydroxipropyl-β-cyclodextrin in aqueous solution. At different times after administration (0.5, and 3 hours), mice were sacrificed (4 animals in each group and at each time) . Blood samples were collected and sera were separated. Antimony concentration was determined in serum by ETAAS with graphite furnace. The results obtained are presented in Table 6. Table 6. Antimony concentrations in mouse sera after 5-fold dilution (μg/L) , at different times after oral administration of a single dose (82.5 mg Sb/kg body weight) of meglumine antimoniate (MA) or its association compound with 2-hydroxypropyl-β-cyclodextrin (MA/Hp-β-CD) . The values are given as average ± standard deviation (n = 4 animals at each time) . Time (hours) MA MA/Hp-β-CD 0.5 90+23 113+10 3 72 +29 168 + 47

When compared to MA, the MA/Hp-β-CD association compound, object of the present invention, showed an antimony concentration significantly larger 3 hours after administration by oral route. It can be concluded that the association of the antimony derivative to Hp-β-CD resulted in increased bioavailability of antimony by oral route, showing the great potential of this preparation in the treatment of leishmaniasis by this administration route. Example 8 Efficacy of the association compound between meglumine antimoniate and β-cyclodextrin in experimental model of cutaneous leishmaniasis. Four groups of BALB/c mice (5 animals per group) were infected on the ear with 2 x 10δ Leishmania amazonensis-GFP (Leishmania amazonensis MHOM/BR/75/strain Josefa transfected by the gene expressing the green fluorescence protein) in the promastigote form. The lesion size was measured periodically and expressed as the difference in thickness between the infected and the non-infected ears. Treatment was started 10 days after infection with the following preparations: the meglumine antimoniate/β- cyclodextrin association compound administered by oral route at a dose of 32 mg Sb/kg (MA/β-CD group) ; meglumine antimoniate in free form administered by oral route at a dose of 120 mg Sb/kg (MA oral group); meglumine antimoniate in free form administered by intraperitoneal at a dose of 60 mg Sb/kg (MA ip group) . The control group received saline by intraperitoneal route. Treatment was carried out on a daily basis, on days 10 to 16 and on days 31 to 36 after infection. Lesion follow-up results are presented in Table 7 for the different groups. Table 7. Evolution of lesion size in BALB/c mice infected with Leishmania amazonensis after treatment with MA/β-CD oral (32 mg Sb/kg) , MA intraperitoneal (60 mg Sb/kg) , MA oral (120 mg Sb/kg) and saline solution.

Lesion size (mm x 10) ± SD Days 5 ii 14 20 28 30 35 40 52 60 68 80 d d d d d d d d d d d d Saline 10 11 14 18 18 22 32 33 39 43 52 66.3 .0 .0 .7 .5 .5 .0 .9 .3 .8 .0 .0 ± 10 ± ± ± ± ± ± ± ± ± ± ± 6. 5. 6. 6. 6. 4. 6. 2. 2. 6. 0. 0 0 0 0 0 9 6 5 5 3 8 MA ip 7. 7. 5. 6. 7. 8. 11 12 13 11 12 18.0 6 4 6 6 2 0 .1 .0 .0 .8 .9 + 0.9 ± ± + ± ± ± ± ± ± ± ± 2. 2. 1. 2. 1. 3. 1. 1. 1. 1. 1. 6 0 8 3 4 0 0 2 3 8 5 MA 7. 8. 9. 27 26 28 29 33 35 38 44 60.8 oral 3 5 8 .8 .7 .3 .1 .2 .8 .0 .6 ± 8.0 ± ± ± ± ± ± ± ± ± ± ± 4. 4. 2. 15 16 3. 6. 6. 9. 12 15 0 0 5 5 5 0 1 MA/β- 6. 6. 5. 10 9. 11 13 11 11 11 16 16.5 CD 0 3 3 .0 0 .8 .0 .0 .1 .7 .5 ± 0.8 oral ± ± ± ± ± ± + + ± + ± 1. 1. 2. 1. 1. 0. 0. 1. 1. 2. 1. 0 5 0 7 0 8 9 1 1 0 2 These results show that the group treated with meglumine antimoniate associated to β-cyclodextrin by oral route presented significantly smaller lesions, when compared to the control group (treated with saline) . On the other hand, the group treated orally with meglumine antimoniate in free form showed similar lesion sizes, when compared to the control group. One can conclude that the association compound is active by oral route against cutaneous leishmaniasis. The leishmanicidal activity of this compound by oral route was equivalent to that observed for the administration of the antimonial in free form given by parenteral (intraperitoneal) route at a twofold higher antimony dose. In addition, the activity of the association compound was significantly higher than that of the antimonial in free form given at a fourfold higher antimony dose. The leishmanicidal activity of the MA/β-CD association compound was also confirmed by the evaluation of the number of parasites present in the lesions. Determination was carried out on day 80 after infection, indirectly, by measuring the fluorescence due to Leishmania amazonensis- GFP parasites. The results of fluorescence measurements, obtained with the different groups, are presented in Table 8. Table 8. Parasite loads in the different mice groups, 80 days after infection. Fluorescence intensity is proportional to the number of parasites. Group Fluorescence + SD

Saline 1.01 ± 0.10 MA ip 0.12 ± 0.07 MA oral 0.74 + 0.09 MA/β-CD oral 0.21 ± 0.03

The data presented in Table 8 show that the parasite load in the group orally treated with the MA/β-CD association compound was significantly lower than that found in the control group and in the group treated with free antimonial by oral route. These data clearly establish that the association of meglumine antimoniate to β-cyclodextrin makes the antimonial active by oral route against cutaneous leishmaniasis. It is important to note that it is the first time that an antimonial composition, showing activity by oral route against cutaneous leishmaniasis, is described in the state of the art.