Synthesis and antileishmanial activity of new 1-aryl-1H-pyrazole-4-carboximidamides derivatives

SHORT REPORT

Synthesis and antileishmanial activity of new 1-aryl-1H-pyrazole-4-carboximidamides derivatives

Maurício S. dos Santos^I; Adriana O. Gomes^I; Alice M. R. Bernardino^I,*; Marcos C. de Souza^I; Misbahul A. Khan^II; Monique A. de Brito^III; Helena C. Castro^IV Paula A. Abreu^IV Carlos R. Rodrigues^V; Rosa M. M. de Léo^VI; Leonor L. Leon^VI; Marilene M. Canto-Cavalheiro^VI

^IPrograma de Pós-Graduação em Química Orgânica and LABioMol, GCM - IB, Universidade Federal Fluminense, Outeiro de São João Baptista, 24020-150 Niterói-RJ, Brazil

^IIChemistry Department, The Islamia University of Bahawalpur, 63100 Bahawalpur, Pakistan

^IIILaboratório de Química Medicinal Computacional, Faculdade de Farmácia, Universidade Federal Fluminense, 24241-000 Niterói-RJ, Brazil

^IVLABioMol, GCM - IB, Universidade Federal Fluminense, Outeiro de São João Baptista, 24020-150 Niterói-RJ, Brazil

^VFaculdade de Farmácia, ModMolQSAR, Universidade Federal do Rio de Janeiro, 24020-150 Rio de Janeiro-RJ, Brazil

^VILaboratório de Bioquímica de Tripanosomatídeos, IOC, Fundação Oswaldo Cruz, 21040-900 Rio de Janeiro-RJ, Brazil

ABSTRACT

Chemotherapy for leishmaniasis, diseases caused by protozoa of the genus Leishmania, remains inefficient in several treatments. So there is a need to search for new drugs. In this work, we have synthesized 1-aryl-1H-pyrazole-4-carboximidamides derivatives and evaluated antileishmanial activities in vitro, as well as cytotoxic effects. Structure-activity relationship (SAR) studies were carried out with all the compounds of the series. Compound 2 showed an activity profile that can be improved through medicinal chemistry strategies.

Keywords: synthesis, 1-aryl-1H-pyrazole-4-carboximidamides, leishmaniasis

RESUMO

A quimioterapia para as leishmanioses, doenças causadas por protozoários do gênero Leishmania, ainda permanece ineficiente em diversos tratamentos. Portanto, existe a necessidade de pesquisa por novos fármacos. Nesse trabalho, foram sintetizados derivados 1-aril-1H-pirazol-4-carboximidamidas, avaliadas as atividades leishmanicida e os efeitos citotóxicos in vitro, e realizado um estudo de relação estrutura-atividade (REA) com a série de compostos. O composto 2 apresentou um perfil de atividade que pode ser melhorado através de estratégias de modificação molecular da química medicinal.

Introduction

Leishmaniasis is a group of vector-borne diseases caused by species of the genus Leishmania that affects about 12 million people in 88 countries in the world. These life-threatening diseases are of medical, social and economic importance in endemic areas, particularly in subtropical and tropical regions. Leishmania parasites exist in two forms: amastigote in the mammalian host and a flagellated promastigote in the insect vector. ^1,2 Clinical manifestations occur in four major forms in humans including: i) visceral leishmaniasis (VL) that is usually fatal when untreated, ii) muco-cutaneous leishmaniasis (MCL) that is a mutilating disease, iii) diffuse cutaneous leishmaniasis (DCL), which is a long-lasting disease due to a deficient cellular-mediated immune response and, iv) cutaneous leishmaniasis (CL) that is disabling when there are multiple lesions. ³

The difficulty to control this parasitic disease remains a serious problem mainly due to the diversity of mammalian reservoirs (wild and domestic animals), species of vectors and Leishmania species. ⁴

Chemotherapy for leishmaniasis is generally ineffective mainly due to the emergence of drug-resistant strains and toxicity of the therapeutics agents. The pentavalent antimonials compounds are widely used as primary therapy whereas other drugs such as amphotericin B, pentamidine, paromomycin, azole derivatives and glucantime are also used. ⁵

Pyrazoles are a class of heterocyclic compounds that exhibit a broad spectrum of biological activities such antiinflamatory, antimicrobial and antitumor. ⁶ Consequently, a large number of synthetic routes to pyrazoles have been reported and summarized in some monographs and reviews. ^7,8 These reports have been useful for biologists and chemists engaged in the development of new drugs and/or synthetic routes. Our group has synthesized pyrazole carbohydrazides with anti-Leishmania in vitro⁹ and in vivo¹⁰ activity. Simultaneously, substances containing the amidine group and affecting large number of pathogens (i.e., Giardia lamblia, Leishmania sp., Pneumocystis carinii, Candida albicans, Aspargillus sp. and Trypanosoma sp. ) have been reported,¹¹ as well as some reviews about synthetic approaches. ¹² Pentamidine (Figure 1) is clinically used in the treatment of pneumonia caused by the opportunistic fungus, Pneumocystis jirovecii, early stage human african tripanosomiasis (HAT) and when treatment with pentavalent antimonials or amphotericin B has failed against Leishmania. ^5,13

In the present work, we prepared five new 1-aryl-1H-pyrazole-4-carboximidamides derivatives 1-5 and evaluated their leishmanicidal acitivity, cytotoxicity and theoretical profiles (Scheme 1).

Results and Discussion

The key intermediates 1-aryl-1H-pyrazole-4-carbonitriles 24-28 were obtained from two alternate routes (methods A and B). In method A, 1-aryl-1H-pyrazole-4-carbaldehydes 10-13 were synthesized through a Vilsmeyer-Haack reaction involving 1-aryl-1H-pyrazoles 6-9, dimethylformamide (DMF) and POCl₃. ¹⁴ The aldehydes formed were converted to key intermediates from the "one-pot reaction" with hydroxylamine and methanoic acid. ¹⁵ In method B, arylhydrazine hydrochlorides 14-18 reacted with ethoxymethylenemalononitrile in ethanol and the resulting 5-amino-1-aryl-1H-pyrazole-4-carbonitriles 19-23 were subjected to aprotic deamination with t-butyl nitrite in tetrahydrofuran to generate the key intermediates. ^16,17 Finally, the targets 1-aryl-1H-pyrazole-4-carboximidamides derivatives 1-5 were obtained by the reaction of the key intermediates 24-28 with gaseous hydrochloric acid followed by treatment with ammonia. ¹⁸

The compounds 1-5 were identified by proton nuclear magnetic resonance (¹H NMR), carbon nuclear magnetic resonance (¹³C NMR), Fourier transform infrared (FTIR) spectroscopies and elementary analysis.

Biological and cytotoxity assays

The effect of different concentrations (40, 80, 160 and 320 µg mL^-1) of the 1-aryl-1H-pyrazole-4-carboximidamides derivatives against L. amazonensis promastigotes growth inhibition was monitored microscopically at the end of the exponential growth phase. Interestingly, an antiproliferative effect was observed for compounds 2 and 3 (IC₅₀ = 105 ± 45 µmol L^-1 or 279 ± 12 µg mL^-1 and 112 ± 41 µmol L^-1 or 259 ± 12 µg mL^-1 respectively) on L .amazonensis in contrast to compounds 1 (IC₅₀ > 1720 µmol L^-1 or > 320 µg mL^-1), 4 (IC₅₀ > 1480 µmol L^-1 or > 320 µg mL^-1) and 5 (IC₅₀ > 1250 µmol L^-1 or > 320 µg mL^-1) (Figure 2). Despite the lower profile of 2 and 3 compared to pentamidine effect (IC₅₀ = 3.6 ± 1.6 µmol L^-1), their activity is still promising since new substitutions may be performed to improve it. The major pyrazolic compounds presented a cytotoxicity profile better than pentamidine (Figure 2).

These biological results may suggest that the two amidine groups of the pentamidine (Figure 1) are important for binding in a non-intercalative way to the minor grove regions of DNA (kDNA) in the Leishmania. ¹⁹

Literature reports described monoamidine derivatives with three hydrophobic phenyl groups with potential effects against L. amazonensis. ²⁰ This effect may be associated to an increase of lipophilicity of these compounds, which facilitates the transport through parasite membrane. In addition, it has been demonstrated the inhibitory effect of these monoamidine derivatives on the phosphorylating activity of cAMP-dependent (cyclic adenosine monophosphate) protein kinase (PKA)²¹ and on nitric oxide production by promastigotes and axenic amastigotes forms of L. amazonensis. ²²

Concerning the 1-aryl-1H-pyrazole-4-carboximidamides compounds, the pyrazoles nucleus have displayed an impressive array of biological activities, among which antiprotozoa, anti-malarial, anti-inflammatory, immunomodulatory, nitric oxide inhibition, cytotoxic and anti-cancer activities. ²³

Molecular modeling and Lipinski rule of five studies

The minimum energy conformations of 1-5 derivatives, calculated by the AM1 semiempirical Hamiltonian,²⁴ showed that, as expected, all rings of these compounds are co-planar except for 5 where the two chlorines lead to a rotation of the ring (Figure 3). Despite of this conformational difference, this structural feature did not contribute for a biological activity for 5. Subsequently, a single-point energy ab initio calculation was performed at the 6-311G* level in order to derive electronic properties, such as highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) energy values, volume, molecular dipole moment (µ), and molecular electrostatic potential (MEP), which could be related to the variation of the antileishmanial activity of these compounds. ²⁵ The results showed that, although the substitution pattern into the 1-aryl-1H-pyrazole-4-carboximidamides structures lead to different antileishmanial profiles, the molecular dipole moment, volume and TPSA (total polar surface area) values did not present any direct correlation with it, as shown in Table 1 and Figure 3.

Differently, HOMO and LUMO energy values apparently seem to be related to these derivatives biological profile since compounds 2 and 3 showed an antileishmanial profile and also presented the lowest values for these orbital energies (Table 1). This suggests a different reactivity for these molecules compared to the other derivatives that probably allowed their active profile.

As receptors recognize stereo-electronic effects and not atom per se, studies of molecular electronic properties could be very effective in interpreting the electronic structure in a comprehensive way. ²⁶ Therefore, the MEP is a useful approach for understanding the electrostatic contribution for the receptor-ligand binding process that has been used in different reports for elucidating this issue. In this work, the analysis of R position on MEPs generated for these compounds revealed that the substitution lead to a different electrostatic distribution depending on the added group mainly on compound 2 (Figure 3). However this MEP difference is not directly expressed in the biological activity as expected.

Since the compounds are considered for oral delivery, in this work we submitted them to the analysis of Lipinski rule of five,²⁷ which indicates if a chemical compound could be an orally active drug in humans. The rule states that most "drug-like" molecules have clogP < 5, molecular weight (M_w) < 500, PSA < 140, and number of hydrogen bond acceptors (HBA) < 10 and donors (HBD) < 5. Molecules violating more than one of these rules may have problems with bioavailability. ²⁷ Our results pointed all compounds as fulfilling this rule and therefore with a good theoretical biodisponibility (Table 1).

Experimental

Unless otherwise noted, all the reagents and solvents were obtained from the market and used without further purification. Melting points were obtained with a Fisher apparatus and were uncorrected. ¹H NMR spectra were recorded at room temperature on a Varian Unity plus 300 MHz employing tetramethylsilane as the internal reference. The chemical shifts (δ) are reported in ppm and the coupling constants (J) in hertz. Infrared (IR) spectra were recorded as potassium bromide (KBr) pellets on a Perkin-Elmer Model 1420 FT IR Spectrophotometer. Microanalyses were performed on a Perkin-Elmer Model 2400 instrument and all values were within ± 0.4% of the calculated compositions. Purity of the reaction products were checked by means of thin layer chromatography (TLC) using silica gel plates with fluorescent indicator and hexane/ethyl acetate (1:1, v/v) as eluent, melting points, IR and ¹H NMR spectra.

General procedure for the preparation of 1-aryl-1H-pyrazole-4-carbaldehydes compounds 10-13

POCl₃ (0.023 mol) was added to DMF (0.033 mol) at 0 ºC and the mixture was stirred for 15 min. After this time 1-aryl-1H-pyrazoles 6-9 (0.003 mol) dissolved in DMF were added dropwise with stirring. The reaction mixture was then heated 3 h at 110 ºC. The solution was then poured slowly into 5 mL saturated sodium carbonate aqueous solution and stirred 30 min. The organic layer was diluted with ether, washed with saturated Na₂CO₃ aqueous solution, and dried with MgSO₄ anhydrous. Evaporation of the organic extract under reduced pressure gave the corresponding products. The recrystallization was made from ethanol/water. 10 (R = H): mp 84-85 ºC, yield 73%; 11 (R = 4-Br): mp 124-126 ºC, yield 86%; 12 (R = 4-NO₂): mp 150-151 ºC, yield 75%; 13 (R = 4-OCH₃): mp 91-92 ºC, yield 59%.

General procedure for the preparation of 5-amino-1-aryl-1H-pyrazole-4-carbonitriles compounds 19-23

Arylhydrazine hydrochlorides 14-18 (0.01 mol) were reacted with ethoxymethylenemalononitrile (0.01 mol) and sodium acetate (0.02 mol) in ethanol (40 mL), under reflux, during 40 min. Afterwards, the mixture was poured in cold water and the precipitate formed was filtered out and recrystallized from ethanol/water. The reactions were accomplished by means of TLC using silica gel plate with fluorescent indicator and hexane/ ethyl acetate (1:1) as eluent. 19 (R=H): mp 135-136 ºC, yield 90%; 20 (R=4-Br): mp 168-169 ºC, yield 86%; 21 (R=4-NO₂): mp 220-221 ºC, yield 68%; 22 (R=4-OCH₃): mp 135-136 ºC, yield 75%; 23 (R=2,6-diCl): mp 190-191 ºC, yield 78%.

General procedure for the preparation of 1-aryl-1H-pyrazole-4-carbonitriles compounds 24-28

Method A

The reaction mixture of the aldehydes 10-13 (7.8 mmol L^-1) with hidroxylamine and methanoic acid (10 mL) was maintained under reflux form 6 h, until the end of reaction was indicated by TLC. Then the reaction mixture was poured in cold water and the precipitate formed was filtered out washed with ethanol and recrystallized from ethanol/water to afford crystals. The purity of the compounds was checked by means of TLC using silica gel plate with fluorescent indicator and hexane/ethyl acetate (1:1, v/v) as eluent, melting point, IR spectra and ¹H NMR. 24 (R = H): mp 91-92 ºC, yield 85%; 25 (R = 4-Br): mp 198-199 ºC, yield 80%; 26 (R = 4-NO₂): mp 185-186 ºC, yield 65%; 27 (R = 4-OCH₃): mp 138-141 ºC, yield 79%; 28 (R = 2,6-diCl): not synthesized.

Method B

The reaction mixture of t-butyl nitrite (4 mL) with dry THF (10 mL) was stirred and refluxed under 20 min. Then, 0.005 mol of 5-amino-1-aryl-1H-pyrazole-4-carbonitriles 19-23 was added. The mixture was stirred and refluxed about 2 h. Afterwards, the mixture THF and t-butyl nitrite was evaporated. The precipitate was recrystallized with the mixture of ethanol/water. The purity of the compounds was checked by means of TLC using silica gel plate with fluorescent indicator and hexane/ethyl acetate (1:1, v/v) as eluent, melting point, IR spectra and ¹H NMR. 24 (R = H): mp 90-91 ºC, yield 82%; 25 (R = 4-Br): mp 198-200 ºC, yield 86%; 26 (R = 4-NO₂): mp 187-188 ºC, yield 62%; 27 (R = 4-OCH₃): mp 138-141 ºC, yield 82%; 28 (R = 2,6-diCl): mp 141-142 ºC, yield 81%.

General procedure for the preparation of 1-aryl-1H-pyrazole-4-carboximidamides compounds 1-5

A mixture of derivatives 24-28 (0.01 mol) and 20 mL of dry ethanol was cooled at 0-5 ºC and saturated with chloridric acid gas. The mixture was sealed and stirred at room temperature for 5 days. After this, bubble ammonium gas was added at mixture reaction and stirred for 7 days. The solvent was evaporated and the crystals was purify with ethanol/water. The purity of the compounds was checked by means of TLC using silica gel plate with fluorescent indicator and hexane/ethyl acetate (1:3) as eluent, melting point, IR Spectra and ¹H NMR.

1-Phenyl-1H-pyrazole-4-carboximidamide (1)

mp 220-221 ºC, yield 67%; FT IR (KBr) ν_max/cm^-1 3303 to 3077, 1682; ¹H NMR (DMSO, δ in ppm): H₃ 8.63 (s), H₅ 9.64 (s), H₂' and H₆' 7.93 (d; 8.1 Hz), H₃' and H₅' 7.71 (t; 7.8 Hz), H₄' 7.56 (t; 7.8 Hz), NHNH₂ 4.08 (br); ¹³C NMR (DMSO, δ in ppm): C₃ 141.3, C₄ 113.3, C₅ 131.4, C₁' 138.7, C₂' and C₆' 119.3, C₃'and C₅' 130.1, C₄' 128.0, C(NH)NH₂ 158.1; MS: m/z 186.2022 (M⁺, 100%). Found: C, 64.34; H, 5.28; N, 29.95. Calc. for C₁₀H₁₀N₄: C, 64.50; H, 5.41; N, 30.09%.

1-(4'-Bromophenyl)-1H-pyrazole-4-carboximidamide (2)

mp 270-271 ºC, yield 66%; FT IR (KBr) ν_max/cm^-1 3400 to 3096, 1660; ¹H NMR (DMSO, δ in ppm): H₃ 9.23 (s), H₅ 10.00 (s), H₂' and H₆' 7.93 (d; 8.7 Hz), H₃' and H₅' 7.21 (d; 9.0 Hz), NHNH₂ 3.48 (br); ¹³C NMR (DMSO, δ in ppm): C₃ 143.1, C₄ 114.2, C₅ 132.5, C₁' 145.8, C₂' and C₆' 124.1, C₃' and C₅' 125.0, C₄' 142.5, C(NH)NH₂ 152.0; MS: m/z 265.1107 (M⁺, 100%). Found: C, 45.21; H, 3.29; N, 21.06; Br, 29.98. Calc. for C₁₀H₉N₄Br: C, 45.30; H, 3.42; N, 21.14; Br, 30.14%.

1-(4'-Nitrophenyl)-1H-pyrazole-4-carboximidamide (3)

mp 257-258 ºC, yield 48%; FT IR (KBr) ν_max/cm^-1 3400 to 3100, 1658; ¹H NMR (DMSO, δ in ppm): H₃ 8.53 (s), H₅ 9.26 (s), 7.20-7.40 (m), NHNH₂ 3.86 (br); ¹³C NMR (DMSO, δ in ppm): C₃ 145.1, C₄ 117.5, C₅ 133.2, C₁' 147.7, C₂' and C₆' 126.0, C₃'and C₅' 127.1, C₄' 146.5, C(NH)NH₂ 156.0; MS: m/z 231.2101 (M⁺, 100%). Found: C, 51.81; H, 3.79; N, 30.06; O, 14.34. Calc. for C₁₀H₉N₅O₂: C, 51.95; H, 3.92; N, 30.29; O, 13.84%.

1-(4'-Methoxyphenyl)-1H-pyrazole-4-carboximidamide (4)

mp 239-240 ºC, yield 65%; FT IR (KBr) ν_max/cm^-1 3211 to 3100, 1634; ¹H NMR (DMSO, δ in ppm): H₃ 8.06 (s), H₅ 8.77 (s), H₂' and H₆' 7.74 (d; 9.0 Hz), H₃' and H₅' 7.06 (d; 8.7 Hz), NHNH₂ 4.62 (br), OCH₃ 3.88 (s); ¹³C NMR (DMSO, δ in ppm): C₃ 143.5, C₄ 92.6, C₅ 120.4, C₁' 131.2, C₂' and C₆' 121.1, C₃'and C₅' 114.9, C₄' 149.8, C(NH)NH₂ 158.8, OCH₃ 55.7; MS: m/z 216.2193 (M⁺, 100%). Found: C, 60.96; H, 5.49; N, 25.75; O, 7.80. Calc. for C₁₁H₁₂N₄O: C, 61.10; H, 5.59; N, 25.91; O, 7.40%.

1-(2',6'-Dichlorophenyl)-1H-pyrazole-4-carboximidamide (5)

mp 222-223 ºC, yield 60%; FT IR (KBr) ν_max/cm^-1 3400 to 3189, 1647; ¹H NMR (DMSO, δ in ppm): H₃ 8.28 (s), H₅ 9.05 (s), H₃' and H₅' 7.93 (d; 8.7 Hz), H₄' 7.55 (t; 8.7 Hz), NHNH₂ 4.65 (br); ¹³C NMR (DMSO, δ in ppm): C₃ 144.0, C₄ 95.9, C₅ 132.1, C₁' 136.8, C₂' 112.4, C₃' and C₅' 118.3, C₄' 128.3, C₅' 130.1, C₆' 112.4, C(NH)NH₂ 156.3; MS: m/z 255.0925 (M⁺, 100%). Found: C, 46.94; H, 3.02; N, 21.88; Cl, 28.16. Calc. for C₁₀H₈N₄Cl₂: C, 47.09; H, 3.16; N, 21.96; Cl, 27.79%.

Biological and cytotoxity assays

Leishmania amazonensis (MHOM/BR/LTB0016 strain) promastigotes were grown at 26 ºC in Schneider' Drosophila medium²¹ supplemented with 10% v/v heat-inactivated foetal calf serum (FCS) at pH 7.2. Parasites were harvested from the medium on day 4, when there was a high percentage of infective forms (metacyclic promastigotes), were counted in a Neubauer' Chamber and adjusted to a concentration of 4×0⁶ parasites mL^-1, for the drug assay. ^22,28

The assay was carried out in 96-well flat-bottom microplate with a volume of 200 mL/well. The compounds 1-5 solubilized in dimethyl sulfoxide (DMSO) (the highest concentration used was 1.6% v/v, not hazardous to the parasite) were added to the culture, in a concentration range from 320 from 80 mg mL^-1. After 24 h incubation in a temperature of 26 ºC, the remaining parasites were counted in a Neubauer's chamber and compared with the controls with DMSO, without the drugs and with the parasites alone. All tests were done in triplicate and pentamidine isethionate was used as reference drug. The IC₅₀/24 h was calculated by means of dose-response curves at a wider range of concentrations, and the results were expressed as the mean ± standard deviation determined from three independent experiments.

The cytotoxicity effect of the derivatives 1-5 expressed as cell viability was assayed on mice's peritoneal macrophages. The cells were isolated from peritoneal cavity of Balb/c mice with cold RPMI 1640 medium, supplemented with 1 mmol L^-1 L-glutamine, 1 mol L^-1 HEPES, penicillin G (10⁵IUI^-1), streptomycin sulfate(0.10 g L^-1). The 2×10⁵ cells per well were cultivated on microplate and incubated at 37 ºC in a humidified 5% CO₂ atmosphere. After 2 h of incubation no adherent cells were then removed and the adhered macrophages were washed twice with RPMI. Compounds were added to the cell culture at the respective EC₅₀ /24 h for L. amazonensis and cells incubated for 24 h. Then, the 3-[4,5-dimethylthiazol-2-yl] -2,5-diphenyl-tetrazolium bromide, MTT was added and after 2-4 h the reaction was interrupted with DMSO. The results could be read in spectrophotometer with wavelength of 570 nm. ^23,28,29

Molecular modeling

The molecular modeling study was performed using SPARTAN'06 software package (Wavefunction Inc. Irvine, CA, 2000). ³⁰ The minimum energy conformation of the derivatives was obtained by the AM1 semiempirical Hamiltonian. In order to better evaluate the electronic properties of the AM1 minimum energy conformations, they were submitted to a single-point energy ab initio calculation at the 6-311G* level.

In order to perform structure-activity relationship (SAR) studies, some electronic properties, such as HOMO and LUMO energy values, HOMO and LUMO orbital coefficients distribution, molecular dipole moment (µ), and molecular electrostatic potential (MEP) were calculated. MEP isoenergy surface maps were generated in the range from -25.0 (deepest red color) to +30.0 (deepest blue color) kcal mol^-1 and superimposed onto a molecular surface of constant electron density of 0.002 e au^-3. Each point of the three dimensional molecular surface map expresses the electrostatic interaction energy value evaluated with a probe atom of positive unitary charge providing an indication of the overall molecular size and location of attractive (negative) or repulsive (positive) electrostatic potentials.

Since the compounds are considered for oral delivery, they were also submitted to the analysis of Lipinski rule of five, which evaluate some properties of a compound that would make it a likely orally active drug in humans. These structural parameters were performed using Molispiration program. ³¹

Conclusions

In this work we described a new set of 1-aryl-1H-pyrazole-4-carboximidamide compounds synthesized in good yields that presented an antileishmanial activity profile. This series can be scaled up and easily produce new analogues. Compound 2 (Br-substituted) also presented a low cytotoxicity profile that pointed it as a lead compound for further substitutions to improve its biological profile. All compounds showed a good theoretical biodisponibility and the molecular modelling evaluation showed that HOMO and LUMO energies of 2 and 3 led to a different reactivity profile that seem to be related to their antileishmanial profile. The hydrophobic substituents in phenyl-pyrazolic groups may be useful to investigate the contribution of this structural unit on its bioactivity profile. Further experiments are being carried out in order to define better chemical structure and biological activity relationships.

Acknowledgments

We thank the Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ), Programa de Desenvolvimento Tecnológico em Insumos para Saúde (PDTIS), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Coordenação de Aperfeiçoamento de Pessoal Docente (CAPES) and Universidade Federal Fluminense (PROPP-UFF) for fellowships and financial support.

References

1. Robert, J. M. H.; Sabourin, C.; Alvarez, N.; Piessard, S. R.; Le Baut, G.; Le Pape, P.; Eur. J. Med. Chem. 2003, 38, 711.

2. Santos, D. O.; Coutinho, C. E. R.; Madeira, M. F.; Bottino, C. G.; Vieira, R. T.; Nascimento, S. B.; Bernardino, A. M. R.; Bourguignon, S. C.; Corte-Real, S.; Pinho, R. T.; Castro, H. C.; Rodrigues, C. R.; Parasitol. Res. 2008, 103, 1.

3. Desjeux, P.; Comp. Immun. Microbiol. Infect. Dis. 2004, 27, 305.

4. Lainson, R.; Shaw, J. J.; The Leishmaniasis in Biology and Medicine, Academic Press: London, 1987.

5. Mayence, A.; Eynde, J. J. V.; LeCour, L.; Walker, L. A.; Tekwani, B. L.; Huang, T. L.; Eur. J. Med. Chem. 2004, 39, 547; Graebin, C. S.; Uchoa, F. D.; Bernardes, L. S. C.; Campo, V. L.; Carvalho, I.; Eifler-Lima, V. L.; Anti-Infect. Agents Med. Chem. 2009, 8, 345; Cavalli, A.; Bolognesi, M. L.; J. Med. Chem. 2009, 52, 7339; Chappuis, F.; Sundar, S.; Hailu, A.; Ghalib, H.; Rijal, S.; Peeling, R. W.; Alvar, J.; Boelaert, M.; Nat. Rev. Microbiol. 2007, 5, 873; Coler, R. N.; Reed, S. G.; Trends Parasitol. 2005, 21, 244.

6. Fancelli, D.; Moll, J.; Varasi, M.; Bravo, R.; Artico, R.; Berta, D.; Bindi, S.; Cameron, A.; Candiani, I.; Cappella, P.; Carpinelli, P.; Croci, W.; Forte, B.; Giorgini, M. L.; Klapwijk, J.; Marsiglio, A.; Pesenti, E.; Rocchetti, M.; Roletto, F.; Severino, D.; Soncini, C.; Storici, P.; Tonani, R.; Zugnoni, P.; Vianello, P.; J. Med. Chem. 2006, 49, 7247; Bekhit, A. A.; Ashour, H. M. A.; Ghany, Y. S. A.; Bekhit, A. E. A.; Baraka, A.; Eur. J. Med. Chem. 2008, 43, 456.

7. Borges, J. C.; Oliveira, C. D.; Pinheiro, L. C. S.; Marra, R. K. F.; Khan, M. A.; Wardell, J. L.; Wardell, S. M. S. V.; Bernardino, A. M. R.; J. Braz. Chem. Soc. 2007, 18, 1571; Bonacorso, H. G.; Oliveira, M. R.; Costa, M. P.; Silva, L. B.; Zanatta, N.; Martins, M. A. B.; Flores, A. F. C.; J. Braz. Chem. Soc. 2005, 16, 868.

8. Makino, K.; Kim, H. S.; Kurasawa, Y.; J. Heterocycl. Chem. 1998, 35, 489.

9. Bernardino, A. M.; Gomes, A. O.; Charret, K. S.; Freitas, A. C.; Machado, G. M.; Canto-Cavalheiro, M. M.; Leon, L. L.; Amaral, V. F.; Eur. J. Med. Chem. 2006, 41, 80.

10. Charret, K. S.; Rodrigues, R. F.; Bernardino, A. M. R.; Gomes, A. O.; Carvalho, A. V.; Canto-Cavalheiro, M. M.; Leon, L. L.; Amaral, V. F.; Am. J. Trop. Med. Hyg. 2009, 80, 568.

11. Croft, S. L.; Coombs, G. H.; Trends Parasitol. 2003, 19, 502; Bernardino, A. M. R.; Gomes, A. O.; Charret, K. S.; Freitas, A. C. C.; Machado, G. M. C.; Canto-Cavalheiro, M. M.; Leon, L. L.; Amaral, V. F.; Eur. J. Med. Chem. 2006, 41, 80; Bell, C. A.; Hall, J. E.; Kyle, D. E.; Antimicrob. Agents Chemother. 1990, 34, 1381; Dardonville, C.; Brun, R.; J. Med. Chem. 2004, 47, 2296; Kramp, K. L.; DeWitt, K.; Flora, J. W.; Muddiman, D. C.; Slunt, K. M.; Houston, T. A.; Tetrahedron Lett. 2005, 46, 695.

12. Santos, M. S.; Bernardino, A. M. R.; Souza, M. C.; Quim. Nova 2006, 29, 1301.

13. Huang, T. L.; Eynde, J. J. V.; Mayence, A.; Collins, M. S.; Cushion, M. T.; Rattendi, D.; Londono, I.; Mazumder, L.; Bacchi, C. J.; Yarlett, N.; Bioorg. Med. Chem. Lett. 2009, 19, 5884.

14. Finar, I. L.; Lord, G. H.; J. Chem. Soc. 1957, 3314.

15. Bekhit, A. A.; Fahmy, H. T. Y.; Rostom, S. A. F.; Baraka, A. M.; Eur. J. Med. Chem. 2003, 38, 27.

16. Sakya, S. M.; Rast, B.; Tetrahedron Lett. 2003, 44, 7629.

17. Cadogan, J. I. G.; Molina, G. A.; J. Chem. Soc. Perkin Trans. I 1973, 541.

18. Tidwell, R. R.; Jones, S. K.; Geratz, J. D.; Ohemeng, K. A.; Cory, M.; Hall, J. E.; J. Med. Chem. 1990, 33, 1252.

19. Bray, P. G.; Barret, M. P.; Ward, S. A.; Koning, H. P.; Trends Parasitol. 2003, 19, 232.

20. Canto-Cavalheiro, M. M.; Echevarria, A.; Araújo, C. A. C.; Bravo, M. F.; Santos, L. H. S.; Jansen, A. M.; Leon, L. L.; Microbios 1997, 90, 51.

21. Genestra, M. J.; Echevarria, A.; Cysne-Finkelstein, L.; Silva-Gonçalves, A. J.; Leon, L. L.; Arzneimittel-Forshung Drug Res. 2001, 51, 920.

22. Temporal, R. M.; Cysne-Finkelstein, L.; Echevarria, A.; Silva-Gonçalves, A. J.; Leon, L. L.; Genestra M. J.; J. Enzyme Inhib. Med. Chem. 2005, 20, 13.

23. Bhat, B. A.; Dhar, K. L.; Puri, S. C.; Saxena, A. K.; Shanmugavel, L. M.; Qazi, G. N.; Bioorg. Med. Chem. Lett. 2005, 15, 3177.

24. Dewar, M. J. S.; Zoebisch, E. G.; Healy, E. F.; Stewart, J. J. P.; J. Am. Chem. Soc. 1985, 107, 3902.

25. Leach, A. R.; Molecular Modeling: Principles and applications, 2^nd ed., Pearson Education: London, 1996.

26. Kubinyi, H.; Pharm. Acta Helv. 1995, 69, 259.

27. Lipinski, C. A.; Lombardo, F.; Dominy, B. W.; Feeney, P. J.; Adv. Drug Deliv. Rev. 2001, 46, 3.

28. Cysne-Finkelstein, L.; Aguiar-Alves, F.; Temporal, R. M.; Leon, L. L.; Exp. Parasitol. 1998, 89, 58.

29. Hendricks, L. D.; Wood, D. E.; Hajduk, M. E.; Parasitology 1978, 76, 309.

30. SPARTAN'06 software package. Wavefunction Inc. Irvine, CA, 2000.

31. Molinspiration Property Calculation Service, www.molinspiration.com.

Submitted: April 28, 2010

Published online: September 21, 2010

References

• 1. Robert, J. M. H.; Sabourin, C.; Alvarez, N.; Piessard, S. R.; Le Baut, G.; Le Pape, P.; Eur. J. Med. Chem. 2003, 38, 711.
• 2. Santos, D. O.; Coutinho, C. E. R.; Madeira, M. F.; Bottino, C. G.; Vieira, R. T.; Nascimento, S. B.; Bernardino, A. M. R.; Bourguignon, S. C.; Corte-Real, S.; Pinho, R. T.; Castro, H. C.; Rodrigues, C. R.; Parasitol. Res. 2008, 103, 1.
• 3. Desjeux, P.; Comp. Immun. Microbiol. Infect. Dis. 2004, 27, 305.
• 4. Lainson, R.; Shaw, J. J.; The Leishmaniasis in Biology and Medicine, Academic Press: London, 1987.
• 5. Mayence, A.; Eynde, J. J. V.; LeCour, L.; Walker, L. A.; Tekwani, B. L.; Huang, T. L.; Eur. J. Med. Chem. 2004, 39, 547;
• Graebin, C. S.; Uchoa, F. D.; Bernardes, L. S. C.; Campo, V. L.; Carvalho, I.; Eifler-Lima, V. L.; Anti-Infect. Agents Med. Chem. 2009, 8, 345;
• Cavalli, A.; Bolognesi, M. L.; J. Med. Chem. 2009, 52, 7339;
• Chappuis, F.; Sundar, S.; Hailu, A.; Ghalib, H.; Rijal, S.; Peeling, R. W.; Alvar, J.; Boelaert, M.; Nat. Rev. Microbiol. 2007, 5, 873;
• Coler, R. N.; Reed, S. G.; Trends Parasitol. 2005, 21, 244.
• 6. Fancelli, D.; Moll, J.; Varasi, M.; Bravo, R.; Artico, R.; Berta, D.; Bindi, S.; Cameron, A.; Candiani, I.; Cappella, P.; Carpinelli, P.; Croci, W.; Forte, B.; Giorgini, M. L.; Klapwijk, J.; Marsiglio, A.; Pesenti, E.; Rocchetti, M.; Roletto, F.; Severino, D.; Soncini, C.; Storici, P.; Tonani, R.; Zugnoni, P.; Vianello, P.; J. Med. Chem. 2006, 49, 7247;
• Bekhit, A. A.; Ashour, H. M. A.; Ghany, Y. S. A.; Bekhit, A. E. A.; Baraka, A.; Eur. J. Med. Chem. 2008, 43, 456.
• 7. Borges, J. C.; Oliveira, C. D.; Pinheiro, L. C. S.; Marra, R. K. F.; Khan, M. A.; Wardell, J. L.; Wardell, S. M. S. V.; Bernardino, A. M. R.; J. Braz. Chem. Soc. 2007, 18, 1571;
• Bonacorso, H. G.; Oliveira, M. R.; Costa, M. P.; Silva, L. B.; Zanatta, N.; Martins, M. A. B.; Flores, A. F. C.; J. Braz. Chem. Soc. 2005, 16, 868.
• 8. Makino, K.; Kim, H. S.; Kurasawa, Y.; J. Heterocycl. Chem. 1998, 35, 489.
• 9. Bernardino, A. M.; Gomes, A. O.; Charret, K. S.; Freitas, A. C.; Machado, G. M.; Canto-Cavalheiro, M. M.; Leon, L. L.; Amaral, V. F.; Eur. J. Med. Chem. 2006, 41, 80.
• 10. Charret, K. S.; Rodrigues, R. F.; Bernardino, A. M. R.; Gomes, A. O.; Carvalho, A. V.; Canto-Cavalheiro, M. M.; Leon, L. L.; Amaral, V. F.; Am. J. Trop. Med. Hyg. 2009, 80, 568.
• 11. Croft, S. L.; Coombs, G. H.; Trends Parasitol 2003, 19, 502;
• Bernardino, A. M. R.; Gomes, A. O.; Charret, K. S.; Freitas, A. C. C.; Machado, G. M. C.; Canto-Cavalheiro, M. M.; Leon, L. L.; Amaral, V. F.; Eur. J. Med. Chem. 2006, 41, 80;
• Bell, C. A.; Hall, J. E.; Kyle, D. E.; Antimicrob. Agents Chemother 1990, 34, 1381;
• Dardonville, C.; Brun, R.; J. Med. Chem. 2004, 47, 2296;
• Kramp, K. L.; DeWitt, K.; Flora, J. W.; Muddiman, D. C.; Slunt, K. M.; Houston, T. A.; Tetrahedron Lett. 2005, 46, 695.
• 12. Santos, M. S.; Bernardino, A. M. R.; Souza, M. C.; Quim. Nova 2006, 29, 1301.
• 13. Huang, T. L.; Eynde, J. J. V.; Mayence, A.; Collins, M. S.; Cushion, M. T.; Rattendi, D.; Londono, I.; Mazumder, L.; Bacchi, C. J.; Yarlett, N.; Bioorg. Med. Chem. Lett. 2009, 19, 5884.
• 14. Finar, I. L.; Lord, G. H.; J. Chem. Soc. 1957, 3314.
• 15. Bekhit, A. A.; Fahmy, H. T. Y.; Rostom, S. A. F.; Baraka, A. M.; Eur. J. Med. Chem. 2003, 38, 27.
• 16. Sakya, S. M.; Rast, B.; Tetrahedron Lett. 2003, 44, 7629.
• 17. Cadogan, J. I. G.; Molina, G. A.; J. Chem. Soc. Perkin Trans. I 1973, 541.
• 18. Tidwell, R. R.; Jones, S. K.; Geratz, J. D.; Ohemeng, K. A.; Cory, M.; Hall, J. E.; J. Med. Chem. 1990, 33, 1252.
• 19. Bray, P. G.; Barret, M. P.; Ward, S. A.; Koning, H. P.; Trends Parasitol. 2003, 19, 232.
• 20. Canto-Cavalheiro, M. M.; Echevarria, A.; Araújo, C. A. C.; Bravo, M. F.; Santos, L. H. S.; Jansen, A. M.; Leon, L. L.; Microbios 1997, 90, 51.
• 21. Genestra, M. J.; Echevarria, A.; Cysne-Finkelstein, L.; Silva-Gonçalves, A. J.; Leon, L. L.; Arzneimittel-Forshung Drug Res. 2001, 51, 920.
• 22. Temporal, R. M.; Cysne-Finkelstein, L.; Echevarria, A.; Silva-Gonçalves, A. J.; Leon, L. L.; Genestra M. J.; J. Enzyme Inhib. Med. Chem. 2005, 20, 13.
• 23. Bhat, B. A.; Dhar, K. L.; Puri, S. C.; Saxena, A. K.; Shanmugavel, L. M.; Qazi, G. N.; Bioorg. Med. Chem. Lett. 2005, 15, 3177.
• 24. Dewar, M. J. S.; Zoebisch, E. G.; Healy, E. F.; Stewart, J. J. P.; J. Am. Chem. Soc. 1985, 107, 3902.
• 25. Leach, A. R.; Molecular Modeling: Principles and applications, 2^nd ed., Pearson Education: London, 1996.
• 26. Kubinyi, H.; Pharm. Acta Helv 1995, 69, 259.
• 27. Lipinski, C. A.; Lombardo, F.; Dominy, B. W.; Feeney, P. J.; Adv. Drug Deliv. Rev. 2001, 46, 3.
• 28. Cysne-Finkelstein, L.; Aguiar-Alves, F.; Temporal, R. M.; Leon, L. L.; Exp. Parasitol. 1998, 89, 58.
• 29. Hendricks, L. D.; Wood, D. E.; Hajduk, M. E.; Parasitology 1978, 76, 309.
• 30. SPARTAN'06 software package. Wavefunction Inc. Irvine, CA, 2000.
• 31. Molinspiration Property Calculation Service, www.molinspiration.com

Publication Dates

• Publication in this collection
  14 Feb 2011
• Date of issue
  Feb 2011

History

• Received
  28 Apr 2010
• Accepted
  21 Sept 2010