Stimulus (polyphenol, IFN-gamma, LPS)-dependent nitric oxide production and antileishmanial effects in RAW 264.7 macrophages

The effects of interferon (IFN-γ), lipopolysaccharide (LPS), and some polyphenols as individual stimuli, as well as in various combinations on NO production in non-infected and infected macrophage-like RAW 264.7 cells were investigated, with emphasis on the NO/parasite kill relationship. In non-infected and in Leishmania parasitized cells, gallic acid significantly inhibited the IFN-γ and LPS-induced NO detected in the supernatant. This effect was less prominent in IFN-γ- than in LPS-stimulated cells. Interestingly, and in contrast to non-infected cells, gallic acid inhibited NO production only when added within 3 h after IFN-γ + LPS. Addition of gallic acid following prolonged incubation with IFN-γ + LPS periods (24 h) no longer inhibited, sometimes even enhanced NO release. Notably, an excellent NO/parasite kill relationship was evident from all the experiments. This study was extended to a series of polyphenols (3-O-shikimic acid, its 3,5-digalloylated analogue, catechin, EGCG, and a procyanidin hexamer) with proven immunostimulatory activities. Although these compounds themselves were found to be weak NO-inducers, the viability of intracellular Leishmania parasites was considerably reduced. Furthermore, their dose-dependent effects on macrophage NO release was determined in the presence of IFN-γ and/or LPS. Again, non-infected and infected cells differed significantly in the NO response, while inhibition of IFN-γ and/or LPS-induced NO production by the tested polyphenols strongly depended on the given time of exposure and the sequence of immunological stimuli. A strong inverse correlation between NO levels and intracellular survival rates of Leishmania parasites supported the assumption that the observed inhibition of NO was not simply due to interference with the Griess assay used for detection.     

The antitumoral, trypanocidal and antileishmanial activities of extract and alkaloids isolated from Duguetia furfuracea

The alkaloid extract and five alkaloids isolated from subterranean stem bark of Duguetia furfuracea (Annonaceae) were investigated for the following activities: antitumoral, trypanocidal and leishmanicidal. Dicentrinone showed weak cytotoxicity, but it had the strongest leishmanicidal activity (IC₅₀ 0.01 μM). Duguetine and duguetine β-N-oxide caused considerable antitumoral activity in every cell lines evaluated, although duguetine was more active against trypomastigote forms (IC₅₀ 9.32 μM) than other alkaloids tested.     

Leishmania (Viannia) panamensis: An in vitro assay using the expression of GFP for screening of antileishmanial drug

Promastigotes of Leishmania (Viannia) panamensis were successfully transfected with p6.5-egfp to express green fluorescent protein. The transfectants remained infective to macrophages, providing an in vitro model for screening antileishmanial drugs. This was demonstrated by flow cytometry of macrophage-associated GFP after exposure of infected cultures to known antileishmanial drugs, i.e. amphotericin B and glucantime^®. Fluorescence of GFP diminished progressively from infected cells with increasing drug concentrations used in both cases. The availability of this fluorescent assay for infection of macrophages by L. (V.) panamensis facilitates drug discovery program for the Viannia species, which differ significantly from those of the Leishmania subgenus.     

Synthesis,antileishmanial activity and docking study of N′-substitutedbenzylidene-2-(6,7-dihydrothieno[3,2-c]pyridin-5(4H)-yl)acetohydrazides

A series of N′-substitutedbenzylidene-2-(6,7-dihydrothieno[3,2-c]pyridin-5(4H)-yl)acetohydrazide derivatives is synthesized and evaluated for antileishmanial activity against Leishmania donovani promastigotes. Compounds 9a and 9i were shown significant antileishmanial when compared with standard sodium stilbogluconate. Antimicrobial study revealed that compound 9b has potent as well as broad spectrum antibacterial activity when compared with ampicillin and compound 9e showed promising antifungal activity when compared with miconazole. Also, none of the synthesized compounds showed cytotoxicity up to tested concentration. Further, docking study against pteridine reductase 1 enzyme of L. donovani showed good binding interactions. ADME properties of synthesized compounds were also analyzed and showed potential to develop as good oral drug candidates.     

Nigrosphaerin A a new isochromene derivative from the endophytic fungus Nigrospora sphaerica

Nigrosphaerin A, a new isochromene derivative (1), was isolated from the endophytic fungus Nigrospora sphaerica and chemically identified as 3-(3,4-dihydroxyphenyl)-4,6,8-trihydroxy-1H-isochromen-1-one-6-O-β-d-glucopyranoside. In addition nineteen known compounds (2–20) were isolated from the same fungus and chemically identified. Compounds (1–3, 5, and 7–16) were isolated for the first time from this fungus. In vitro antileukemic, antileishmanial, antifungal, antibacterial and antimalarial activities of (1–20) were examined. Compounds 5, 7, 9 and 10 showed good antileukemic activity against HL60 cells with IC₅₀ values of 0.03, 0.39, 0.2 and 0.4 μg/mL, respectively and against K562 cells with IC₅₀ values of 0.35, 0.35, 0.49 and 0.01 μg/mL, respectively. Compounds 3, 4 and 6 showed moderate antileishmanial activity with IC₅₀ values of 30.2, 26.4 and 36.4 μg/ml, respectively. Compound 7 showed moderate antifungal activity against Cryptococcus neoformans with IC₅₀ value of 14.8 μg/mL.     

Microwave-assisted synthesis of novel 5-substituted benzylidene amino-2-butyl benzofuran-3-yl-4-methoxyphenyl methanones as antileishmanial and antioxidant agents

A series of 5-substitutedbenzylideneamino-2-butylbenzofuran-3-yl-4-methoxyphenyl methanones is synthesized and evaluated for antileishmanial and antioxidant activities. Compounds 4f (IC₅₀?=?52.0?±?0.09?µg/ml), 4h (IC₅₀?=?56.0?±?0.71?µg/ml) and 4l (IC₅₀?=?59.3?±?0.55?µg/ml) were shown significant antileishmanial when compared with standard sodium stibogluconate (IC₅₀?=?490.0?±?1.5?µg/ml). Antioxidant study revealed that compounds 4i (IC₅₀?=?2.44?±?0.47?µg/ml) and 4l (IC₅₀?=?3.69?±?0.44?µg/ml) have shown potent comparable activity when compared with standard ascorbic acid (IC₅₀?=?3.31?±?0.34?µg/ml). Molecular docking study was carried out which replicating results of biological activity in case of initial hits 4f and 4h suggesting that these compounds have a potential to become lead molecules in drug discovery process. In silico ADME study was performed for predicting pharmacokinetic profile of the synthesised antileishmanial agents and expressed good oral drug like behaviour.     

New series of 3,5-disubstituted tetrahydro-2H-1,3,5-thiadiazine thione (THTT) derivatives: Synthesis and potent antileishmanial activity

Four series of heterocyclic compounds, namely, tetrahydro-2H-1,3,5-thiadiazine thione derivatives were synthesized in good to excellent yields and were screened for their in vitro antileishmanial activities against Leishmania major (promastigotes). Most of the compounds showed significant antileishmanial activity within the range of IC₅₀?=?15.48–39.36?μM when compared with standard pentamidine (IC₅₀?=?14.95?μM). The structure-activity relationship showed that N-3 and N-5 substituents have a key role against leishmanicidal activity. The ester analogues (series B) were found to have a 1.5 to 5-fold reduced activity compared to their acidic counterparts. Cytotoxicity against mammalian mouse fibroblast 3?T3 cells was also evaluated and compared between the acid and its ester analogue. The reduction of antileishmanial activity and loss of toxicity in the newly developed THTT ester derivative indicates that these compounds can be used as a template study for the production of effective antileishmanial ester prodrugs.     

Synthesis of N-(1-methyl-1H-indol-3-yl)methyleneamines and 3,3-diaryl-4-(1-methyl-1H-indol-3-yl)azetidin-2-ones as potential antileishmanial agents

A series of N-(1-methyl-1H-indol-3-yl)methyleneamines and eight new 3,3-diaryl-4-(1-methyl-1H-indol-3-yl)azetidin-2-ones have been synthesized and screened for their antileishmanial activity against Leishmania major. 3,3-Diaryl-4-(1-methyl-1H-indol-3-yl)azetidin-2-ones have been synthesized by the Staudinger's ketene-imine cycloaddition employing two 2-diazo-1,2-diarylethanones as the precursors of diarylketenes. A marked improvement in anti-parasitic activity is observed by transformation of the methyleneamines to azetidin-2-ones in seven out of eight compounds. Two compounds displayed antileishmanial activity comparable to that of the clinically used antileshmanial drug, amphotericine B.     

Abietane diterpenoids and triterpenoic acids from Salvia cilicica and their antileishmanial activities

Bioguided-fractionation of an acetone extract of the roots of Salvia cilicica (Lamiaceae) led to isolation of two new diterpenes, 7-hydroxy-12-methoxy-20-nor-abieta-1,5(10),7,9,12-pentaen-6,14-dione and abieta-8,12-dien-11,14-dione (12-deoxy-royleanone), together with oleanolic acid, ursolic acid, ferruginol, inuroyoleanol and cryptanol. Their structures were determined spectroscopically, which included HREIMS and 2D NMR spectroscopic analysis. The new abietane derivatives showed appreciable in vitro antileishmanial activity against intracellular amastigote forms of both Leishmania donovani (IC(50) values of 170 and 120 nM, respectively) and Leishmania major (IC(50) values of 290 and 180 nM, respectively). The triterpenoic acids were found to be potently active against amastigote (IC(50) values of 7-120 nM) and moderately active against promastigote stages (IC(50) values of 51-137 nM) of the two Leishmania species.     

Chemical constituents from Baphia leptobotrys Harms (Fabaceae) and their chemophenetic significance

The chemical investigation of the CH₂Cl₂/MeOH (1:1) extract of the trunk bark and leaves of Baphia leptobotrys Harms (Fabaceae) led to the isolation of nineteen compounds (1–19). Their structures were elucidated based on the analysis of their NMR and MS data. Except for compounds 3–5, 14 and 15, all the isolated compounds are reported from the genus Baphia for the first time. Furthermore, compounds 8, 10, 16, 17 are isolated from the Fabaceae family for the first time. The crude extract and isolated compounds were screened in vitro for their antileishmanial activity against Leishmania donovani 1S (MHOM/SD/62/1S) promastigotes and cytotoxicity towards Raw 264.7 macrophage cells. The crude extract of B. leptobotrys exhibited moderate activity (IC₅₀ = 63.40 μg/mL). Amongst the tested compounds, ergosterol peroxide (9) and germanicol caffeoyl ester (8) exhibited good activity with IC₅₀ values of 9.43 and 10.24 μM, respectively and showed acceptable selectivity towards macrophage cells (SI > 3.85). The chemophenetic significance of these compounds is also discussed.