Natural terpenoids from Ambrosia species are active in vitro and in vivo against human pathogenic trypanosomatids

Among the natural compounds, terpenoids play an important role in the drug discovery process for tropical diseases. The aim of the present work was to isolate antiprotozoal compounds from Ambrosia elatior and A. scabra. The sesquiterpene lactone (STL) cumanin was isolated from A. elatior whereas two other STLs, psilostachyin and cordilin, and one sterol glycoside, daucosterol, were isolated from A. scabra. Cumanin and cordilin were active against Trypanosoma cruzi epimastigotes showing 50% inhibition concentrations (IC50) values of 12 µM and 26 µM, respectively. Moreover, these compounds are active against bloodstrean trypomastigotes, regardless of the T. cruzi strain tested. Psilostachyin and cumanin were also active against amastigote forms with IC50 values of 21 µM and 8 µM, respectively. By contrast, daucosterol showed moderate activity on epimastigotes and trypomastigotes and was inactive against amastigote forms. We also found that cumanin and psilostachyin exhibited an additive effect in their trypanocidal activity when these two drugs were tested together. Cumanin has leishmanicidal activity with growth inhibition values greater than 80% at a concentration of 5 µg/ml (19 µM), against both L. braziliensis and L. amazonensis promastigotes. In an in vivo model of T. cruzi infection, cumanin was more active than benznidazole, producing an 8-fold reduction in parasitemia levels during the acute phase of the infection compared with the control group, and more importantly, a reduction in mortality with 66% of the animals surviving, in comparison with 100% mortality in the control group. Cumanin also showed nontoxic effects at the doses assayed in vivo, as determined using markers of hepatic damage.Fil: Sülsen, Valeria P.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Metabolismo del Fármaco (i); ArgentinaFil: Cazorla, Silvia Ines. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Oficina de Coordinacion Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral "Profesor R. A. Margni"; ArgentinaFil: Frank, Fernanda Maria. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Oficina de Coordinacion Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral "Profesor R. A. Margni"; ArgentinaFil: Laurella, Laura C.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Metabolismo del Fármaco (i); ArgentinaFil: Muschietti, Liliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Metabolismo del Fármaco (i); ArgentinaFil: Catalan, Cesar Atilio Nazareno. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Tucumán. Instituto de Quimica del Noroeste; ArgentinaFil: Martino, Virginia Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Metabolismo del Fármaco (i); ArgentinaFil: Malchiodi, Emilio Luis. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Oficina de Coordinacion Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral "Profesor R. A. Margni"; Argentin

South American medicinal flora: a promising source of novel compounds with antiprotozoal activity

Native populations of South America have employed herb-based preparations for the treatment of parasite diseases. In this review, some examples of South American medicinal plants from which bioactive molecules have been isolated are presented. Results of our research related to the study of novel compounds with antiprotozoal activity are also presented herein. Peruvin and psilostachyin, two sesquiterpene lactones isolated from the Argentine medicinal species Ambrosia tenuifolia, presented significative in vitro activity on Trypanosoma cruzi epimastigotes and trypomastigotes. Psilostachyin also presented in vivo activity in T. cruzi infected mice. Both compounds were also active on Leishmania spp. The results obtained suggest that psilostachyin could be considered a potential lead molecule in the development of novel trypanocidal agents.Colegio de Farmacéuticos de la Provincia de Buenos Aire

Búsqueda de compuestos antiprotozoarios en especies de la flora medicinal argentina

Los extractos orgánicos y acuosos de once especies medicinales argentinas fueron evaluados in vitro sobre epimastigotes de Trypanosoma cruzi. Las especies Ambrosia tenuifolia, A. scabra, Baccharis spicata, Acanthostyles buniifolium (Asteraceae), Lippia integrifolia (Verbenaceae) y Clinopodium gilliesii (Lamiaceae) fueron activas con inhibiciones mayores al 70 % en la concentración de 100 µg/ml. Los extractos orgánicos de A. tenuifolia y de A. scabra presentaron además, actividades leishmanicida y antiplasmódica con porcentajes de inhibición superiores al 50 y 40 %, respectivamente, a 10 µg/ml. Del fraccionamiento bioguiado del extracto orgánico de A. tenuifolia se aislaron dos lactonas sesquiterpénicas y un flavonoide activos sobre epimastigotes de T. cruzi: psilostachina (IC50 = 1,2 µg/ml) y peruvina (IC50 = 1,6 µg/ml) e hispidulina (IC50 = 14,0 µg/ml), respectivamente. A partir del extracto orgánico de A. scabra se aisló la lactona sesquiterpénica psilostachina C, que fue activa con un valor de IC50 de 0,6 µg/ml. La psilostachina y la psilostachina C también fueron activas in vitro sobre tripomastigotes de T. cruzi, promastigotes de Leishmania mexicana y sobre cepas sensibles y resistentes a cloroquina de Plasmodium falciparum. Ambos compuestos presentaron actividad in vivo sobre ratones infectados con T. cruzi, disminuyeron el número de parásitos circulantes y aumentaron el tiempo de sobrevida de los animales. Psilostachina y psilostachina C indujeron alteraciones ultraestructurales en T. cruzi a concentraciones menores que 1 µg/ml

Mode of Action of the Sesquiterpene Lactones Psilostachyin and Psilostachyin C on Trypanosoma cruzi.

Trypanosoma cruzi is the causative agent of Chagas' disease, which is a major endemic disease in Latin America and is recognized by the WHO as one of the 17 neglected tropical diseases in the world. Psilostachyin and psilostachyin C, two sesquiterpene lactones isolated from Ambrosia spp., have been demonstrated to have trypanocidal activity. Considering both the potential therapeutic targets present in the parasite, and the several mechanisms of action proposed for sesquiterpene lactones, the aim of this work was to characterize the mode of action of psilostachyin and psilostachyin C on Trypanosoma cruzi and to identify the possible targets for these molecules. Psilostachyin and psilostachyin C were isolated from Ambrosia tenuifolia and Ambrosia scabra, respectively. Interaction of sesquiterpene lactones with hemin, the induction of oxidative stress, the inhibition of cruzipain and trypanothione reductase and their ability to inhibit sterol biosynthesis were evaluated. The induction of cell death by apoptosis was also evaluated by analyzing phosphatidylserine exposure detected using annexin-V/propidium iodide, decreased mitochondrial membrane potential, assessed with Rhodamine 123 and nuclear DNA fragmentation evaluated by the TUNEL assay. Both STLs were capable of interacting with hemin. Psilostachyin increased about 5 times the generation of reactive oxygen species in Trypanosoma cruzi after a 4h treatment, unlike psilostachyin C which induced an increase in reactive oxygen species levels of only 1.5 times. Only psilostachyin C was able to inhibit the biosynthesis of ergosterol, causing an accumulation of squalene. Both sesquiterpene lactones induced parasite death by apoptosis. Upon evaluating the combination of both compounds, and additive trypanocidal effect was observed. Despite their structural similarity, both sesquiterpene lactones exerted their anti-T. cruzi activity through interaction with different targets. Psilostachyin accomplished its antiparasitic effect by interacting with hemin, while psilostachyin C interfered with sterol synthesis

Natural Sesquiterpene Lactones Induce Oxidative Stress in Leishmania mexicana

Leishmaniasis is a worldwide parasitic disease, caused by monoflagellate parasites of the genus Leishmania. In the search for more effective agents against these parasites, the identification of molecular targets has been attempted to ensure the efficiency of drugs and to avoid collateral damages on the host's cells. In this work, we have investigated some of the mechanisms of action of a group of natural sesquiterpene lactones that are effective against Leishmania mexicana mexicana promastigotes. We first observed that the antiproliferative effect of mexicanin I (Mxc), dehydroleucodine (DhL), psilostachyin (Psi), and, at lesser extent, psilostachyin C (Psi C) is blocked by 1.5 mM reduced glutathione. The reducing agent was also able to reverse the early effect of the compounds, suggesting that lactones may react with intracellular sulfhydryl groups. Moreover, we have shown that all the sesquiterpene lactones, except Psi C, significantly decreased the endogenous concentration of glutathione within the parasite. Consistent with these findings, the active sesquiterpene lactones increased between 2.7 and 5.4 times the generation of ROS by parasites. These results indicate that the induction of oxidative stress is at least one of the mechanisms of action of DhL, Mxc, and Psi on parasites while Psi C would act by another mechanism

Antiprotozoal Compounds from Urolepis hecatantha (Asteraceae)

The dewaxed dichloromethane extract of Urolepis hecatantha and the compounds isolated from it were tested for their in vitro activity on Trypanosoma cruzi epimastigotes and Leishmania infantum promastigotes. The extract of U. hecatantha showed activity against both parasites with IC50 values of 7 µg/mL and 31 µg/mL, respectively. Fractionation of the dichloromethane extract led to the isolation of euparin, jaceidin, santhemoidin C, and eucannabinolide. The sesquiterpene lactones eucannabinolide and santhemoidin C were active on T. cruzi with IC50 values of 10 ± 2 µM (4.2 µg/mL) and 18 ± 3 µM (7.6 µg/mL), respectively. Euparin and santhemoidin C were the most active on L. infantum with IC50 values of 18 ± 4 µM (3.9 µg/mL) and 19 ± 4 µM (8.0 µg/mL), respectively. Eucannabinolide has also shown drug-like pharmacokinetic and toxicity properties

Effect of Psi and PsiC on cell death.

<p>Figs a-c: <i>T</i>. <i>cruzi</i> epimastigotes were treated with Psi and PsiC 35 μM during 8, 24 and 48 h and stained with Annexin V-FITC/PI (a and b). Bars correspond to AV^-PI^-: viable cells, AV⁺PI^-: early apoptotic cells, AV⁺PI⁺: late apoptotic cells and AV^-PI⁺: necrotic cells. Epimastigotes exposed to 30% fresh human serum (FHS) for 2 h at 28°C were used as positive control. p values < 0.05 (*) and < 0.01 (**) were considered significant.</p

Effect of Psi and PsiC on epimastigotes cultured with different hemin concentrations.

<p><i>T</i>. <i>cruzi</i> epimastigotes were grown at 28°C for 4 days with Psi (a) and PsiC (b) (2.5–15 μM) in culture medium containing different hemin concentrations (0, 5, 10 and 20 mg/L). (c) The IC₅₀ (50% inhibitory concentration) values for each compound with different hemin concentrations were estimated by lineal regression analysis from the inhibition percentage values and the decimal logarithm (log) of drug concentration.</p