Synthesis, molecular docking studies, and larvicidal activity evaluation of new fluorinated neonicotinoids against Anopheles darlingi larvae.

Anopheles darlingi is the main vector of malaria in Brazil, characterized by a high level of anthropophilia and endophagy. Imidacloprid, thiacloprid, and acetamiprid are the most widespread insecticides of the neonicotinoid group. However, they produce adverse effects on the non-target insects. Flupyradifurone has been marketed as an alternative to non-fluorinated neonicotinoids. Neonicotinoids containing trifluoroacethyl substituent reveal increased insecticidal activity due to higher hydrophobicity and metabolic stability. We synthesized novel neonicotinoid insecticides containing fluorinated acceptor groups and their interactions were estimated with the nicotinic acetylcholine receptor (nAChR) binding site by molecular docking studies, to evaluate their larvicidal activity against A. darlingi, and to assess their outdoor photodegradation behavior. New neonicotinoid analogues were prepared and characterized by NMR and mass-spectrometry. The synthesized molecules were modelled by time-dependent density functional theory and analyzed, their interaction with nAChR was investigated by molecular docking. Their insecticide activity was tested on Anopheles larvae collected in suburban area of Manaus, Brazil. Four new fluorinated neonicotinoid analogs were prepared and tested against 3rd instars larvae of A. darlingi showing high larvicidal activity. Docking studies reveal binding modes of the synthesized compounds and suggest that their insecticidal potency is governed by specific interactions with the receptor binding site and enhanced lipophilicity. 2-Chloro-5-(2-trifluoromethyl-pyrrolidin-1-ylmethyl)pyridine 5 showed fast degradation in water maintaining high larvicidal activity. All obtained substances possessed high larvicidal activity in low concentrations in 48 hours of exposure, compared to commercial flupyradifurone. Such activity is connected to a unique binding pattern of the synthesized compounds to insect's nAChR and to their enhanced bioavailability owing to introduction of fluorinated amino-moieties. Therefore, the compounds in question have a high potential for application as control agents for insects transmitting tropical diseases, and they will be less persistent in the environment.

Antileishmanial activity of extracts from Libidibia ferrea: development of in vitro and in vivo tests / Atividade anti-leishmania de extratos de Libidibia ferrea: desenvolvimento de testes in vitro e in vivo

ABSTRACT Treatment of cutaneous leishmaniasis (CL) is difficult due to the scarce number of drugs able to eliminate completely the intracellular form of the parasite. In the present study, the aim was to evaluate: i) phytochemical properties of extracts from Libidibia ferrea; ii) antileishmanial activity of extracts from L. ferrea against promastigotes and amastigotes of Leishmania (Leishmania) amazonensis and Leishmania (Viannia) guyanensis; iii) the effects of topical treatment using hydrogel containing active extract of L. ferrea on golden hamsters infected with L. (L.) amazonensis. Extracts from leaves, branches and fruits of L. ferrea were obtained with hexane and methanol and were tested by in vitro assays in promastigotes and murine macrophages J774 experimentally infected with amastigotes of Leishmania. Groups of hamsters with CL received topical treatment with a formulation of extract (10%) hydrogels, 50 mg.day-1 for 40 days. In vitro activity of FrMeOH (methanolic extract from fruits without seeds) resulted in significant reduction of viable promastigotes of L. (L.) amazonensis (IC50 of 15.4 µg.mL-1) and demonstrated inhibition potential of amastigote forms of L. (L.) amazonensis and L. (V.) guyanensis and low cytotoxicity in macrophages. The overall data of topical treatment with extract hydrogels (GelFrMeOH) showed that lesion sizes were significantly reduced (42.78%), with low parasite burden by RT-qPCR and culture analysis by microscopy examination, and with histopathological findings such as lower inflammatory cell infiltration 40 days after treatment. Chemical analysis demonstrated FrMeOH contains high levels of phenolic compounds. The results indicate a possible alternative therapy for CL using phytotherapics.(AU)

RESUMO O tratamento da leishmaniose cutânea (LC) é de difícil resultado, devido ao escasso número de fármacos capazes de eliminar completamente a forma intracelular do parasita. No presente estudo, objetivou-se avaliar: i) propriedades fitoquímicas dos extratos de Libidibia ferrea; ii) a atividade antileishmania de extratos de L. ferrea contra promastigotas e amastigotas de Leishmania (Leishmania) amazonensis e Leishmania (Viannia) guyanensis; iii) os efeitos do tratamento tópico utilizando hidrogel contendo extrato ativo de L. ferrea em hamsters dourados (Mesocricetus auratus) infectados com L. (L.) amazonensis. Extratos de folhas, galhos e frutos de L. ferrea foram obtidos com hexano e metanol e foram testados por ensaios in vitro contra promastigotas e macrófagos J774 infectados com amastigotas de Leishmania. Grupos de hamsters infectados receberam hidrogel tópico com extrato (10%), 50 mg.dia-1 durante 40 dias. A atividade in vitro de FrMeOH (extrato metanólico dos frutos) mostrou redução significativa de promastigotas de L. (L.) amazonensis (IC50 de 15,4 g.mL-1), potencial de inibição de formas amastigotas de L. (L.) amazonensis e L. (V.) guyanensis (IC50 303,36 g.mL-1) e baixa citotoxicidade em células de macrófagos J774. Os resultados destacaram que as lesões cutâneas de animais que receberam tratamento com hidrogel + extrato (GelFrMeOH) apresentaram redução significativa (42,78%), menos úlceração e redução da carga parasitária detectada por RT-qPCR, microscopia e análise em cultura e alterações histopatológicas, como menor infiltrado inflamatório após 40 dias de tratamento. A análise química demonstrou que FrMeOH contém altos níveis de compostos fenólicos. Os resultados apontam para uma possível terapia alternativa para CL utilizando fitoterápicos.(AU)

Antileishmanial activity of extracts from Libidibia ferrea: development of in vitro and in vivo tests / Atividade anti-leishmania de extratos de Libidibia ferrea: desenvolvimento de testes in vitro e in vivo

ABSTRACT Treatment of cutaneous leishmaniasis (CL) is difficult due to the scarce number of drugs able to eliminate completely the intracellular form of the parasite. In the present study, the aim was to evaluate: i) phytochemical properties of extracts from Libidibia ferrea; ii) antileishmanial activity of extracts from L. ferrea against promastigotes and amastigotes of Leishmania (Leishmania) amazonensis and Leishmania (Viannia) guyanensis; iii) the effects of topical treatment using hydrogel containing active extract of L. ferrea on golden hamsters infected with L. (L.) amazonensis. Extracts from leaves, branches and fruits of L. ferrea were obtained with hexane and methanol and were tested by in vitro assays in promastigotes and murine macrophages J774 experimentally infected with amastigotes of Leishmania. Groups of hamsters with CL received topical treatment with a formulation of extract (10%) hydrogels, 50 mg.day-1 for 40 days. In vitro activity of FrMeOH (methanolic extract from fruits without seeds) resulted in significant reduction of viable promastigotes of L. (L.) amazonensis (IC50 of 15.4 µg.mL-1) and demonstrated inhibition potential of amastigote forms of L. (L.) amazonensis and L. (V.) guyanensis and low cytotoxicity in macrophages. The overall data of topical treatment with extract hydrogels (GelFrMeOH) showed that lesion sizes were significantly reduced (42.78%), with low parasite burden by RT-qPCR and culture analysis by microscopy examination, and with histopathological findings such as lower inflammatory cell infiltration 40 days after treatment. Chemical analysis demonstrated FrMeOH contains high levels of phenolic compounds. The results indicate a possible alternative therapy for CL using phytotherapics.

RESUMO O tratamento da leishmaniose cutânea (LC) é de difícil resultado, devido ao escasso número de fármacos capazes de eliminar completamente a forma intracelular do parasita. No presente estudo, objetivou-se avaliar: i) propriedades fitoquímicas dos extratos de Libidibia ferrea; ii) a atividade antileishmania de extratos de L. ferrea contra promastigotas e amastigotas de Leishmania (Leishmania) amazonensis e Leishmania (Viannia) guyanensis; iii) os efeitos do tratamento tópico utilizando hidrogel contendo extrato ativo de L. ferrea em hamsters dourados (Mesocricetus auratus) infectados com L. (L.) amazonensis. Extratos de folhas, galhos e frutos de L. ferrea foram obtidos com hexano e metanol e foram testados por ensaios in vitro contra promastigotas e macrófagos J774 infectados com amastigotas de Leishmania. Grupos de hamsters infectados receberam hidrogel tópico com extrato (10%), 50 mg.dia-1 durante 40 dias. A atividade in vitro de FrMeOH (extrato metanólico dos frutos) mostrou redução significativa de promastigotas de L. (L.) amazonensis (IC50 de 15,4 μg.mL-1), potencial de inibição de formas amastigotas de L. (L.) amazonensis e L. (V.) guyanensis (IC50 303,36 μg.mL-1) e baixa citotoxicidade em células de macrófagos J774. Os resultados destacaram que as lesões cutâneas de animais que receberam tratamento com hidrogel + extrato (GelFrMeOH) apresentaram redução significativa (42,78%), menos úlceração e redução da carga parasitária detectada por RT-qPCR, microscopia e análise em cultura e alterações histopatológicas, como menor infiltrado inflamatório após 40 dias de tratamento. A análise química demonstrou que FrMeOH contém altos níveis de compostos fenólicos. Os resultados apontam para uma possível terapia alternativa para CL utilizando fitoterápicos.

Topical treatment of experimental cutaneous leishmaniasis in golden hamster (Mesocricetus auratus) with formulations containing pentamidine / Tratamento tópico da leishmaniose cutânea experimental em golden hamsters (Mesocricetus auratus) com fórmulas contendo pentamidina

Current treatment of cutaneous leishmaniasis (CL) relies mainly on pentavalent antimonials salts and second-line drugs include pentamidine and amphotericin B, but these therapies have side effects and require parenteral administration. The aim of this work was to evaluate the topical formulations containing pentamidine isethionate (PI) in the experimental treatment of cutaneous leishmaniasis (CL). Golden hamsters (Mesocricetus auratus) were infected in the nose with Leishmania (Leishmania) amazonensis. Six treatment groups received different topical treatments of anhydrous or hydrating emulsions, for a maximum of 10 days, with an application of 50 mg day-1. After treatment tissue samples of lesions were evaluated by histology, transmission electron microscopy and biopsy cultivation. Compared with untreated group, topical treatment with hydrating emulsion with 10% PI and usnic acid (ACE5AU) showed significantly decrease in volume lesion (P= 0.028) on 20th day after the end of the treatment with reduction of 27.37%. Topical treatment with anhydrous emulsion with 10% PI and usnic acid (ACPU) reduces parasite burden in Golden hamsters. This study demonstrated the potential of topical treatment to reduce the number of parasites that could be combined with others drugs and to have a faster and more effective treatment of cutaneous leishmaniasis.(AU)

O tratamento atual para Leishmaniose Tegumentar Americana é realizado com antimoniais pentavalentes como tratamento padrão, e as drogas de segunda escolha incluem pentamidina e anfotericina B, mas essas terapias apresentam efeitos colaterais e requerem administração parenteral. O objetivo deste trabalho foi avaliar formulações tópicas contendo isetionato de pentamidina comercial (IP) no tratamento da leishmaniose cutânea experimental (LC). Hamsters dourados foram infectados no focinho com Leishmania (Leishmania) amazonensis. Grupos de seis animais receberam tratamento tópico com cremes anidro ou emulsões hidratantes, num máximo de 10 dias, com aplicação de 50 mg dia-1. Amostras de tecido de lesões tratadas foram avaliadas por histologia, microscopia eletrônica de transmissão (MET) e cultivo de biópsia. Comparado ao grupo sem tratamento, o tratamento tópico com emulsão hidratante com 10% de IP e ácido úsnico (ACE5AU) mostrou diminuição significativa (P=0,028) nas medidas de lesões, 20 dias após o final do tratamento e 27,37% de redução. O tratamento tópico com emulsão anidra com 10% de IP e ácido úsnico (ACPU) mostrou redução da carga parasitária em Golden hamsters. Este estudo demonstrou a possibilidade de utilizar o tratamento tópico para reduzir o número de parasitas e que este poderia ser associado a outras drogas para tratamento mais rápido e eficaz da leishmaniose cutânea.(AU)

Topical treatment of experimental cutaneous leishmaniasis in golden hamster (Mesocricetus auratus) with formulations containing pentamidine

O tratamento atual para Leishmaniose Tegumentar Americana é realizado com antimoniais pentavalentes como tratamento padrão, e as drogas de segunda escolha incluem pentamidina e anfotericina B, mas essas terapias apresentam efeitos colaterais e requerem administração parenteral. O objetivo deste trabalho foi avaliar formulações tópicas contendo isetionato de pentamidina comercial (IP) no tratamento da leishmaniose cutânea experimental (LC). Hamsters dourados foram infectados no focinho com Leishmania (Leishmania) amazonensis. Grupos de seis animais receberam tratamento tópico com cremes anidro ou emulsões hidratantes, num máximo de 10 dias, com aplicação de 50 mg dia-1. Amostras de tecido de lesões tratadas foram avaliadas por histologia, microscopia eletrônica de transmissão (MET) e cultivo de biópsia. Comparado ao grupo sem tratamento, o tratamento tópico com emulsão hidratante com 10% de IP e ácido úsnico (ACE5AU) mostrou diminuição significativa (P=0,028) nas medidas de lesões, 20 dias após o final do tratamento e 27,37% de redução. O tratamento tópico com emulsão anidra com 10% de IP e ácido úsnico (ACPU) mostrou redução a carga parasitária em Golden hamsters. Este estudo demonstrou a possibilidade de utilizar o tratamento tópico para reduzir o número de parasitas e que este poderia ser associado a outras drogas para tratamento mais rápido e eficaz da leishmaniose cutânea.(AU)

Nanoscaled hydrated antimony (V) oxide as a new approach to first-line antileishmanial drugs.

BACKGROUND: Coordination compounds of pentavalent antimony have been, and remain, the first-line drugs in leishmaniasis treatment for >70 years. Molecular forms of Sb (V) complexes are commercialized as sodium stibogluconate (Pentostam®) and meglumine antimoniate (MA) (Glucantime®). Ever-increasing drug resistance in the parasites limits the use of antimonials, due to the low drug concentrations being administered against high parasitic counts. Sb5+ toxicity provokes severe side effects during treatment. To enhance therapeutic potency and to increase Sb (V) concentration within the target cells, we decided to try a new active substance form, a hydrosol of Sb2O5·nH2O nanoparticles (NPs), instead of molecular drugs. METHODOLOGY/PRINCIPAL FINDINGS: Sb2O5·nH2O NPs were synthesized by controlled SbCl5 hydrolysis in a great excess of water. Sb2O5·nH2O phase formation was confirmed by X-ray diffraction. The surface of Sb (V) NPs was treated with ligands with a high affinity for target cell membrane receptors. The mean particle size determined by dynamic light scattering and transmission electron microscopy was ~35-45 nm. In vitro tests demonstrated a 2.5-3 times higher antiparasitic activity of Sb (V) nanohybrid hydrosols, when compared to MA solution. A similar comparison for in vivo treatment of experimental cutaneous leishmaniasis with Sb5+ nanohybrids showed a 1.75-1.85 times more effective decrease in the lesions. Microimages of tissue fragments confirmed the presence of NPs inside the cytoplasm of infected macrophages. CONCLUSION/SIGNIFICANCE: Sb2O5·nH2O hydrosols are proposed as a new form of treatment for cutaneous leishmaniasis caused by Leishmania amazonensis. The NPs penetrate directly into the affected cells, creating a high local concentration of the drug, a precondition to overcoming the parasite resistance to molecular forms of pentavalent antimonials. The nanohybrids are more effective at a lower dose, when compared to MA, the molecular drug. Our data suggest that the new form of treatment has the potential to reduce and simplify the course of cutaneous leishmaniasis treatment. At the same time, Sb2O5·nH2O hydrosols provide an opportunity to avoid toxic antimony (V) spreading throughout the body.

Antiplasmodial activity of synthetic ellipticine derivatives and an isolated analog.

Ellipticine has been shown previously to exhibit excellent in vitro antiplasmodial activity and in vivo antimalarial properties that are comparable to those of the control drug chloroquine in a mouse malaria model. Ellipticine derivatives and analogs exhibit antimalarial potential however only a few have been studied to date. Herein, ellipticine and a structural analog were isolated from Aspidosperma vargasii bark. A-ring brominated and nitrated ellipticine derivatives exhibit good in vitro inhibition of Plasmodium falciparum K1 and 3D7 strains. Several of the compounds were found not to be toxic to human fetal lung fibroblasts. 9-Nitroellipticine (IC50=0.55µM) exhibits greater antiplasmodial activity than ellipticine. These results are further evidence of the antimalarial potential of ellipticine derivatives.