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1.
J Ethnopharmacol ; 264: 113262, 2021 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-32818574

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: In the Peruvian Amazon as in the tropical countries of South America, the use of medicinal Piper species (cordoncillos) is common practice, particularly against symptoms of infection by protozoal parasites. However, there is few documented information about the practical aspects of their use and few scientific validation. The starting point of this work was a set of interviews of people living in six rural communities from the Peruvian Amazon (Alto Amazonas Province) about their uses of plants from Piper genus: one community of Amerindian native people (Shawi community) and five communities of mestizos. Infections caused by parasitic protozoa take a huge toll on public health in the Amazonian communities, who partly fight it using traditional remedies. Validation of these traditional practices contributes to public health care efficiency and may help to identify new antiprotozoal compounds. AIMS OF STUDY: To record and validate the use of medicinal Piper species by rural people of Alto Amazonas Province (Peru) and annotate active compounds using a correlation study and a data mining approach. MATERIALS AND METHODS: Rural communities were interviewed about traditional medication against parasite infections with medicinal Piper species. Ethnopharmacological surveys were undertaken in five mestizo villages, namely: Nueva Arica, Shucushuyacu, Parinari, Lagunas and Esperanza, and one Shawi community (Balsapuerto village). All communities belong to the Alto Amazonas Province (Loreto region, Peru). Seventeen Piper species were collected according to their traditional use for the treatment of parasitic diseases, 35 extracts (leaves or leaves and stems) were tested in vitro on P. falciparum (3D7 chloroquine-sensitive strain and W2 chloroquine-resistant strain), Leishmania donovani LV9 strain and Trypanosoma brucei gambiense. Assessments were performed on HUVEC cells and RAW 264.7 macrophages. The annotation of active compounds was realized by metabolomic analysis and molecular networking approach. RESULTS: Nine extracts were active (IC50 ≤ 10 µg/mL) on 3D7 P. falciparum and only one on W2 P. falciparum, six on L. donovani (axenic and intramacrophagic amastigotes) and seven on Trypanosoma brucei gambiense. Only one extract was active on all three parasites (P. lineatum). After metabolomic analyses and annotation of compounds active on Leishmania, P. strigosum and P. pseudoarboreum were considered as potential sources of leishmanicidal compounds. CONCLUSIONS: This ethnopharmacological study and the associated in vitro bioassays corroborated the relevance of use of Piper species in the Amazonian traditional medicine, especially in Peru. A series of Piper species with few previously available phytochemical data have good antiprotozoal activity and could be a starting point for subsequent promising work. Metabolomic approach appears to be a smart, quick but still limited methodology to identify compounds with high probability of biological activity.


Assuntos
Antiprotozoários/metabolismo , Etnofarmacologia/métodos , Medicina Tradicional/métodos , Metabolômica/métodos , Piper/metabolismo , Extratos Vegetais/metabolismo , Animais , Antimaláricos/isolamento & purificação , Antimaláricos/metabolismo , Antimaláricos/uso terapêutico , Antiprotozoários/isolamento & purificação , Antiprotozoários/uso terapêutico , Feminino , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/metabolismo , Mesocricetus , Camundongos , Peru/etnologia , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/uso terapêutico , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/metabolismo , Células RAW 264.7 , Inquéritos e Questionários
2.
PLoS Negl Trop Dis ; 14(8): e0008575, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866156

RESUMO

BACKGROUND: Treatment failure and resistance to the commonly used drugs remains a major obstacle for successful chemotherapy against visceral leishmaniasis (VL). Since the development of novel therapeutics involves exorbitant costs, the effectiveness of the currently available antitrypanosomatid drug suramin has been investigated as an antileishmanial, specifically for VL,in vitro and in animal model experiments. METHODOLOGY/PRINCIPAL: Leishmania donovani promastigotes were treated with suramin and studies were performed to determine the extent and mode of cell mortality, cell cycle arrest and other in vitro parameters. In addition, L. donovani infected BALB/c mice were administered suramin and a host of immunological parameters determined to estimate the antileishmanial potency of the drug. Finally, isothermal titration calorimetry (ITC) and enzymatic assays were used to probe the interaction of the drug with one of its putative targets namely parasitic phosphoglycerate kinase (LmPGK). FINDINGS: The in vitro studies revealed the potential efficacy of suramin against the Leishmania parasite. This observation was further substantiated in the in vivo murine model, which demonstrated that upon suramin administration, the Leishmania infected BALB/c mice were able to reduce the parasitic burden and also generate the host protective immunological responses. ITC and enzyme assays confirmed the binding and consequent inhibition of LmPGK due to the drug. CONCLUSIONS/SIGNIFICANCE: All experiments affirmed the efficacy of suramin against L. donovani infection, which could possibly lead to its inclusion in the repertoire of drugs against VL.


Assuntos
Antiprotozoários/farmacologia , Antiprotozoários/uso terapêutico , Leishmaniose Visceral/tratamento farmacológico , Suramina/farmacologia , Suramina/uso terapêutico , Animais , Apoptose/efeitos dos fármacos , Citocinas/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Concentração Inibidora 50 , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/parasitologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Óxido Nítrico/metabolismo , Fosfoglicerato Quinase/efeitos dos fármacos , Células RAW 264.7/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo
3.
Parasitol Res ; 119(9): 2991-3003, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32748038

RESUMO

Visceral leishmaniasis (VL, also known as kala-azar) is a vector borne disease caused by obligate intracellular protozoan parasite Leishmania donovani. To overcome the limitations of currently available drugs for VL, molecular target-based study is a promising tool to develop new drugs to treat this neglected tropical disease. One such target we recently identified from L. donovani (Ld) genome (WGS, clinical Indian isolate; BHU 1220, AVPQ01000001) is a small GTP-binding protein, Rab6 protein. We now report a specific inhibitor of the GTPase activity of Rab6 protein of L. donovani (LdRab6) without restricting host enzyme activity. First, to understand the nature of LdRab6 protein, we generated recombinant LdRab6 mutant proteins (rLdRab6) by systematically introducing deletion (two cysteine residues at C-terminal) and mutations [single amino acid substitutions in the conserved region of GTP (Q84L)/GDP(T38N) coding sequence]. The GTPase activity of rLdRab6:GTP and rLdRab6:GDP locked mutant proteins showed ~ 8-fold and ~ 1.5-fold decreases in enzyme activity, respectively, compared to the wild type enzyme activity. The mutant protein rLdRab6:ΔC inhibited the GTPase activity. Sequence alignment analysis of Rab6 protein of L. donovani with Homo sapiens showed identical amino acids in the G conserved region (GTP/GDP-binding sites) but it differed in the C-terminal region. We then evaluated the inhibitory activity of trans-dibenzalacetone (DBA, a synthetic analog of curcumin with strong antileishmanial activity reported earlier by us) in the GTPase activity of LdRab6 protein. Comparative molecular docking analysis of DBA and specific inhibitors of Rab proteins (Lovastatin, BFA, Zoledronate, and NE10790) indicated that DBA had optimum binding affinity with LdRab6 protein. This was further confirmed by the GTPase activity of DBA-treated LdRab6 which showed a basal GTP level significantly lower than that of the wild-type rLdRab6. The results confirm that DBA inhibits the GTPase activity of LdRab6 protein from L. donovani (LdRab6), a potential target for its antileishmanial effect.


Assuntos
Antiprotozoários/farmacologia , Inibidores Enzimáticos/farmacologia , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/parasitologia , Pentanonas/farmacologia , Proteínas de Protozoários/antagonistas & inibidores , Proteínas rab de Ligação ao GTP/antagonistas & inibidores , Sequência de Aminoácidos , Sítios de Ligação , Curcumina/farmacologia , Humanos , Leishmania donovani/química , Leishmania donovani/enzimologia , Leishmania donovani/genética , Leishmaniose Visceral/tratamento farmacológico , Simulação de Acoplamento Molecular , Pentanonas/química , Proteínas de Protozoários/química , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Alinhamento de Sequência , Proteínas rab de Ligação ao GTP/química , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo
4.
PLoS One ; 15(7): e0236127, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32692783

RESUMO

Visceral leishmaniasis (VL) is an important zoonosis in Brazil. Dogs are considered the main domestic reservoirs of the disease in the country; hence, control measures are focused on these reservoirs. Despite efforts to prevent and control VL, important reductions in disease prevalence and incidence have not been identified, stimulating the development and application of new strategies. The choice and implementation of new control strategies can benefit from the application of mathematical models that allow the simulation of different strategies in different scenarios. Selecting the best strategy to be implemented is also supported by cost-effectiveness studies. Here we used the results of a mathematical model in which scenarios, including isolated use of the vaccine and insecticide-impregnated collar (IIC), both at different coverage rates, were simulated to conduct a cost-effectiveness study. The costs were calculated for each scenario considering a simulation period of four years. Collar application in both infected and non-infected animals was the most cost-effective strategy. For example, to reduce the prevalence in humans and dogs by approximately 70%, the costs ranged from $250,000 and $550,000 for the IICs and vaccination, respectively. Even in the scenario with 40% loss/replacement of IICs, this measure was more advantageous in terms of cost-effectiveness than vaccination. If the vaccine were applied with culling of seropositive tested dogs, then the measure became more effective with a reduced cost compared with the vaccine alone. The use of the three first consecutive vaccine doses had the greatest impact on the cost of the vaccination strategy. The advantage of using IICs is that there is no need for a prior diagnosis, unlike vaccination, reducing costs and facilitating implementation. The present study aims to contribute to strategies to reduce hosts infected with VL by reducing public expenditure.


Assuntos
Doenças do Cão/economia , Doenças do Cão/prevenção & controle , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/economia , Leishmaniose Visceral/prevenção & controle , Vacinação/economia , Vacinação/veterinária , Animais , Brasil/epidemiologia , Doenças do Cão/parasitologia , Doenças do Cão/transmissão , Cães , Leishmania donovani/isolamento & purificação , Leishmaniose Visceral/epidemiologia , Leishmaniose Visceral/veterinária , Prevalência
5.
Exp Parasitol ; 217: 107948, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32698076

RESUMO

Immunomodulation is an emerging concept to combat infection in recent years. Immunomodulators like arabinosylated-lipoarabinomannan (Ara-LAM) and glycyrrhizic-acid (GA) possess anti-leishmanial property, whereas sodium-antimony-gluconate (SAG) is still considered as the first choice for chemotherapy against leishmaniasis. During infection, invasion of Leishmania donovani needs the potential requirement of Ca2+, which is further responsible for apoptosis in intracellular amastigotes. However, suppression of elevated intracellular calcium by the activation of plasma-membrane-calcium-ATPase (PMCA4) facilitates survival of L. donovani in the host. In the present study, SAG, Ara-LAM, and GA were found to evoke significant increase in intracellular Ca2+ in L. donovani infected macrophages by inhibiting PMCA4. Moreover, PMCA4 inhibition by TFP or PMCA4 siRNA elevated the level of PKCß, whereas calcium-independent upregulation of PKCζ remained unchanged in infected macrophages. Furthermore, application of immunomodulators in infected macrophages resulted in down-regulation of PKCζ, conversion of anti-inflammatory to pro-inflammatory cytokines and inhibition of PMCA4. Plasma membrane-associated ceramide which is known to be elevated during leishmaniasis, triggered upregulation of PMCA4 via PKCζ activation. Interestingly, immunomodulators attenuated ceramide generation, which resulted into reduced PKCζ activation leading to the decreased PMCA expression in infected macrophages. Therefore, our study elucidated the efficacy of SAG, Ara-LAM, and GA in the reduction of parasite burden in macrophages by suppressing PMCA activation through inhibition of ceramide mediated upregulation of PKCζ.


Assuntos
Antiprotozoários/uso terapêutico , ATPases Transportadoras de Cálcio/sangue , Membrana Celular/enzimologia , Fatores Imunológicos/farmacologia , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/tratamento farmacológico , Animais , Gluconato de Antimônio e Sódio/farmacologia , Gluconato de Antimônio e Sódio/uso terapêutico , Antiprotozoários/farmacologia , Cálcio/metabolismo , ATPases Transportadoras de Cálcio/efeitos dos fármacos , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Ceramidas/metabolismo , Meios de Cultura Livres de Soro , Densitometria , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Ácido Glicirrízico/farmacologia , Ácido Glicirrízico/uso terapêutico , Imipramina/farmacologia , Immunoblotting , Lipopolissacarídeos/farmacologia , Lipopolissacarídeos/uso terapêutico , Macrófagos/fisiologia , Camundongos , RNA de Protozoário/genética , RNA de Protozoário/isolamento & purificação , RNA Interferente Pequeno/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transcrição Reversa , Transfecção
6.
Sci Rep ; 10(1): 12243, 2020 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-32699361

RESUMO

The development of an effective oral therapeutics is an immediate need for the control and elimination of visceral leishmaniasis (VL). We exemplify the preparation and optimization of 2-hydroxypropyl-ß-cyclodextrin (HPCD) modified solid lipid nanoparticles (SLNs) based oral combinational cargo system of Amphotericin B (AmB) and Paromomycin (PM) against murine VL. The emulsion solvent evaporation method was employed to prepare HPCD modified dual drug-loaded solid lipid nanoparticles (m-DDSLNs). The optimized formulations have a mean particle size of 141 ± 3.2 nm, a polydispersity index of 0.248 ± 0.11 and entrapment efficiency for AmB and PM was found to be 96% and 90% respectively. The morphology of m-DDSLNs was confirmed by scanning electron microscopy and transmission electron microscopy. The developed formulations revealed a sustained drug release profile upto 57% (AmB) and 21.5% (PM) within 72 h and were stable at both 4 °C and 25 °C during short term stability studies performed for 2 months. Confocal laser scanning microscopy confirmed complete cellular internalization of SLNs within 24 h of incubation. In vitro cytotoxicity study against J774A.1 macrophage cells confirmed the safety and biocompatibility of the developed formulations. Further, m-DDSLNs did not induce any hepatic/renal toxicities in Swiss albino mice. The in vitro simulated study was performed to check the stability in simulated gastric fluids and simulated intestinal fluids and the release was found almost negligible. The in vitro anti-leishmanial activity of m-DDSLNs (1 µg/ml) has shown a maximum percentage of inhibition (96.22%) on intra-cellular amastigote growth of L. donovani. m-DDSLNs (20 mg/kg × 5 days, p.o.) has significantly (P < 0.01) reduced the liver parasite burden as compared to miltefosine (3 mg/kg × 5 days, p.o.) in L. donovani-infected BALB/c mice. This work suggests that the superiority of as-prepared m-DDSLNs as a promising approach towards the oral delivery of anti-leishmanial drugs.


Assuntos
Anfotericina B/química , Anfotericina B/farmacologia , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/tratamento farmacológico , Nanopartículas/química , Paromomicina/química , Paromomicina/farmacologia , Animais , Antiprotozoários/química , Antiprotozoários/farmacologia , Linhagem Celular , Emulsões/química , Lipídeos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Transmissão/métodos , Tamanho da Partícula , Fosforilcolina/análogos & derivados , Fosforilcolina/química , Fosforilcolina/farmacologia
7.
J Med Chem ; 63(8): 4306-4314, 2020 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-32223141

RESUMO

We report for the first time a novel series of tellurides bearing sulfonamide as selective and potent inhibitors of the ß-class carbonic anhydrase (CA; EC 4.2.1.1) enzyme expressed in Leishmania donovani protozoa. Such derivatives showed high activity against axenic amastigotes, and among them, compound 5g (4-(((3,4,5-trimethoxyphenyl)tellanyl)methyl)benzenesulfonamide) showed an IC50 of 0.02 µM being highly selective for the parasites over THP-1 cells with a selectivity index of 300. The in vitro and in vivo toxicity experiments showed compound 5g to possess a safe profile and thus paving the way for tellurium-containing compounds as novel drug entities.


Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Leishmania donovani/efeitos dos fármacos , Sulfonamidas/farmacologia , Tripanossomicidas/farmacologia , Animais , Inibidores da Anidrase Carbônica/química , Relação Dose-Resposta a Droga , Feminino , Jejuno/efeitos dos fármacos , Jejuno/patologia , Leishmania donovani/fisiologia , Técnicas de Cultura de Órgãos , Ratos , Ratos Wistar , Sulfonamidas/química , Tripanossomicidas/química
8.
PLoS Negl Trop Dis ; 14(4): e0008188, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32275665

RESUMO

Leishmaniasis is one of the Neglected Tropical Diseases (NTDs) which is closely associated with poverty and has gained much relevance recently due to its opportunistic coinfection with HIV. It is a protozoan zoonotic disease transmitted by a dipteran Phlebotomus, Lutzomyia/ Sergentomyia sandfly; during blood meals on its vertebrate intermediate hosts. It is a four-faceted disease with its visceral form being more deadly if left untreated. It is endemic across the tropics and sub-tropical regions of the world. It can be considered the third most important NTD after malaria and lymphatic filariasis. Currently, there are numerous drawbacks on the fight against leishmaniasis which includes: non-availability of vaccines, limited availability of drugs, high cost of mainstay drugs and parasite resistance to current treatments. In this study, we screened the antileishmanial activity, selectivity, morphological alterations, cell cycle progression and apoptotic potentials of six Pathogen box compounds from Medicine for Malaria Venture (MMV) against Leishmania donovani promastigotes and amastigotes. From this study, five of the compounds showed great promise as lead chemotherapeutics based on their high selectivity against the Leishmania donovani parasite when tested against the murine mammalian macrophage RAW 264.7 cell line (with a therapeutic index ranging between 19-914 (promastigotes) and 1-453 (amastigotes)). The cell cycle progression showed growth arrest at the G0-G1 phase of mitotic division, with an indication of apoptosis induced by two (2) of the pathogen box compounds tested. Our findings present useful information on the therapeutic potential of these compounds in leishmaniasis. We recommend further in vivo studies on these compounds to substantiate observations made in the in vitro study.


Assuntos
Antiprotozoários/farmacologia , Desenvolvimento de Medicamentos , Leishmania donovani/efeitos dos fármacos , Anfotericina B/farmacologia , Animais , Apoptose/efeitos dos fármacos , Concentração Inibidora 50 , Cinética , Leishmania donovani/crescimento & desenvolvimento , Macrófagos/parasitologia , Camundongos , Microscopia de Fluorescência , Células RAW 264.7
9.
Parasite Immunol ; 42(6): e12719, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32248547

RESUMO

AIMS: Visceral leishmaniasis (VL), caused by Leishmania donovani in India, is fatal if untreated, having serious concern of limited chemotherapeutic options. In this study, we evaluated antileishmanial efficacy of purified chlorogenic acid (CGA) against promastigotes and intracellular amastigotes infected into RAW264.7 macrophages. METHODS AND RESULTS: Chlorogenic acid was effective both on promastigotes (IC50  = 78.394 µmol/L, i.e. 27.75 µg/mL) and intracellular amastigotes (ED50  = 26.752 µmol/L, i.e. 9.47 µg/mL). In promastigotes, significant retardation in mitotic growth was caused both by cell-death and reduction of metabolic activity, evidenced by propidium-iodide uptake and MTT assay, respectively. Flow cytometric analysis revealed that retardation of mitotic growth was due to cell-cycle arrest at G1/S checkpoint. Complete clearance of amastigotes from infected RAW264.7 cells, assessed by microscopic counting, was achieved with 60 µmol/L (21.24 µg/mL) CGA for 24 hours, with negligible toxicity to host macrophages. This parasite clearing efficacy was comparable to 1.0 µg/mL (1.082 µmol/L) Amphotericin B, and 20 µmol/L Miltefosine, two standard antileishmanial drugs. Cytokine-ELISA revealed that elevated IL-10 production by infected macrophages was reduced after parasite clearance. Consequently, IL-12, TNF and NO (assayed by Griess test) production by macrophages were significantly increased after successful resolution of infection. CONCLUSION: Chlorogenic acid might emerge as a potential antileishmanial drug.


Assuntos
Antiprotozoários/uso terapêutico , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Ácido Clorogênico/uso terapêutico , Citocinas/metabolismo , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/tratamento farmacológico , Óxido Nítrico/metabolismo , Animais , Linhagem Celular , Índia , Leishmaniose Visceral/mortalidade , Leishmaniose Visceral/parasitologia , Macrófagos/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Fosforilcolina/análogos & derivados , Fosforilcolina/uso terapêutico , Células RAW 264.7
10.
Parasit Vectors ; 13(1): 96, 2020 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-32087758

RESUMO

BACKGROUND: Since the introduction of miltefosine (MIL) as first-line therapy in the kala-azar elimination programme in the Indian subcontinent, treatment failure rates have been increasing. Since parasite infectivity and virulence may become altered upon treatment relapse, this laboratory study assessed the phenotypic effects of repeated in vitro and in vivo MIL exposure. METHODS: Syngeneic Leishmania donovani lines either or not exposed to MIL were compared for drug susceptibility, rate of promastigote multiplication and metacyclogenesis, macrophage infectivity and behaviour in the sand fly vector, Lutzomyia longipalpis. RESULTS: Promastigotes of both in vitro and in vivo MIL-selected strains displayed a slightly reduced drug susceptibility that was associated with a reduced MIL-accumulation linked to a lower copy number (disomic state) of chromosome 13 harboring the miltefosine transporter (LdMT) gene. In vitro selected promastigotes showed a lower rate of metacyclogenesis whereas the in vivo derived promastigotes displayed a moderately increased growth rate. Repeated MIL exposure did neither influence the parasite load nor metacyclogenesis in the sand fly vector. CONCLUSIONS: Recurrent in vitro and in vivo MIL exposure evokes a number of very subtle phenotypic and genotypic changes which could make promastigotes less susceptible to MIL without attaining full resistance. These changes did not significantly impact on infection in the sand fly vector.


Assuntos
Antiprotozoários/farmacologia , Insetos Vetores/parasitologia , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/fisiologia , Fosforilcolina/análogos & derivados , Psychodidae/parasitologia , Aclimatação , Animais , Resistência a Medicamentos , Humanos , Leishmania donovani/patogenicidade , Leishmaniose Visceral/parasitologia , Leishmaniose Visceral/transmissão , Testes de Sensibilidade Parasitária , Fenótipo , Fosforilcolina/farmacologia , Virulência
11.
Eur J Med Chem ; 191: 112146, 2020 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-32088496

RESUMO

In continuation of our pharmacomodulation work on the nitroimidazooxazole series, we report the synthesis of new 5-substituted 6-nitroimidazooxazole derivatives. Our aim was to evaluate how functionalization of the 5-position of the 6-nitroimidazooxazole scaffold affects antileishmanial and antitrypanosomal in vitro activities. Twenty-one original compounds were synthesized and evaluated for their in vitro antileishmanial (L. donovani) and antitrypanosomal (T. cruzi) properties. Pallado-catalyzed cross-coupling reactions were used to introduce an aryl or ethynyl aryl substituent in 5-position from a 5-brominated-6-nitroimidazooxazole starting product. Unfortunately, the first series of compounds bearing an aryl group in 5-position presented limited in vitro activities against L. donovani and T. cruzi, with IC50 > 10 µM (vs 0.18 µM and 2.31 µM for the reference drugs amphotericin B and benznidazole respectively). Interestingly, the second series of compounds bearing an ethynyl aryl substituent in 5-position showed more promising, particularly against T. cruzi. Compounds 6a, 6b, 6c, 6g and 6h had better activity than the reference drug benznidazole (0.92 µM ≤ IC50 ≤ 2.18 µM vs IC50 = 2.31 µM), whereas the non-functionalized 2-methyl-6-nitro-2,3-dihydroimidazo [2,1-b]oxazole 2 was not active against T. cruzi (IC50 > 10 µM).


Assuntos
Antiprotozoários/farmacologia , Leishmania donovani/efeitos dos fármacos , Nitroimidazóis/farmacologia , Oxazóis/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Antiprotozoários/síntese química , Antiprotozoários/química , Relação Dose-Resposta a Droga , Estrutura Molecular , Nitroimidazóis/síntese química , Nitroimidazóis/química , Oxazóis/síntese química , Oxazóis/química , Testes de Sensibilidade Parasitária , Relação Estrutura-Atividade
12.
Parasit Vectors ; 13(1): 94, 2020 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-32085719

RESUMO

BACKGROUND: New therapeutic drugs are urgently needed against visceral leishmaniasis because current drugs, such as pentavalent antimonials and miltefosine, produce severe side effects and development of resistance. Whether cyclosporine A (CsA) and its derivatives can be used as therapeutic drugs for visceral leishmaniasis has been controversial for many years. METHODS: In this study, we evaluated the efficacy of CsA and its derivative, dihydrocyclosporin A (DHCsA-d), against promastigotes and intracellular amastigotes of Leishmania donovani. Sodium stibogluconate (SSG) was used as a positive control. RESULTS: Our results showed that DHCsA-d was able to inhibit the proliferation of L. donovani promastigotes (IC50: 21.24 µM and 12.14 µM at 24 h and 48 h, respectively) and intracellular amastigotes (IC50: 5.23 µM and 4.84 µM at 24 and 48 h, respectively) in vitro, but CsA treatment increased the number of amastigotes in host cells. Both DHCsA-d and CsA caused several alterations in the morphology and ultrastructure of L. donovani, especially in the mitochondria. However, DHCsA-d showed high cytotoxicity towards cells of the mouse macrophage cell line RAW264.7, with CC50 values of 7.98 µM (24 h) and 6.65 µM (48 h). Moreover, DHCsA-d could increase IL-12, TNF-α and IFN-γ production and decrease the levels of IL-10, IL-4, NO and H2O2 in infected macrophages. On the contrary, CsA decreased IL-12, TNF-α, and IFN-γ production and increased the levels of IL-10, IL-4, NO and H2O2 in infected macrophages. The expression of L. donovani cyclophilin A (LdCyPA) in promastigotes and intracellular amastigotes and the expression of cyclophilin A (CyPA) in RAW 264.7 cells were found to be significantly downregulated in the CsA-treated group compared to those in the untreated group. However, no significant changes in LdCyPA and CyPA levels were found after DHCsA-d or SSG treatment. CONCLUSIONS: Our findings initially resolved the dispute regarding the efficacy of CsA and DHCsA-d for visceral leishmaniasis treatment. CsA showed no significant inhibitory effect on intracellular amastigotes. DHCsA-d significantly inhibited promastigotes and intracellular amastigotes, but it was highly cytotoxic. Therefore, CsA and DHCsA-d are not recommended as antileishmanial drugs.


Assuntos
Antiprotozoários/farmacologia , Ciclosporina/farmacologia , Ciclosporinas/farmacologia , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/parasitologia , Animais , Avaliação Pré-Clínica de Medicamentos , Humanos , Interferon gama/imunologia , Interleucina-10/imunologia , Interleucina-2/imunologia , Leishmania donovani/crescimento & desenvolvimento , Leishmania donovani/fisiologia , Leishmaniose Visceral/tratamento farmacológico , Leishmaniose Visceral/imunologia , Macrófagos/imunologia , Macrófagos/parasitologia , Camundongos , Células RAW 264.7
13.
PLoS Negl Trop Dis ; 14(2): e0007991, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32023254

RESUMO

BACKGROUND: During infections involving intracellular pathogens, iron performs a double-edged function by providing the pathogen with nutrients, but also boosts the host's antimicrobial arsenal. Although the role of iron has been described in visceral leishmaniasis, information regarding its status in the dermal sequel, Post Kala-azar Dermal Leishmaniasis (PKDL) remains limited. Accordingly, this study aimed to establish the status of iron within monocytes/macrophages of PKDL cases. METHODOLOGY/PRINCIPAL FINDINGS: The intramonocytic labile iron pool (LIP), status of CD163 (hemoglobin-haptoglobin scavenging receptor) and CD71 (transferrin receptor, Tfr) were evaluated within CD14+ monocytes by flow cytometry, and soluble CD163 by ELISA. At the lesional sites, Fe3+ status was evaluated by Prussian blue staining, parasite load by qPCR, while the mRNA expression of Tfr (TfR1/CD71), CD163, divalent metal transporter-1 (DMT-1), Lipocalin-2 (Lcn-2), Heme-oxygenase-1 (HO-1), Ferritin, Natural resistance-associated macrophage protein (NRAMP-1) and Ferroportin (Fpn-1) was evaluated by droplet digital PCR. Circulating monocytes demonstrated elevated levels of CD71, CD163 and soluble CD163, which corroborated with an enhanced lesional mRNA expression of TfR, CD163, DMT1 and Lcn-2. Additionally, the LIP was raised along with an elevated mRNA expression of ferritin and HO-1, as also iron exporters NRAMP-1 and Fpn-1. CONCLUSIONS/SIGNIFICANCE: In monocytes/macrophages of PKDL cases, enhancement of the iron influx gateways (TfR, CD163, DMT-1 and Lcn-2) possibly accounted for the enhanced LIP. However, enhancement of the iron exporters (NRAMP-1 and Fpn-1) defied the classical Ferritinlow/Ferroportinhigh phenotype of alternatively activated macrophages. The creation of such a pro-parasitic environment suggests incorporation of chemotherapeutic strategies wherein the availability of iron to the parasite can be restricted.


Assuntos
Ferro/metabolismo , Leishmaniose Cutânea/metabolismo , Adolescente , Adulto , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/genética , Antígenos de Diferenciação Mielomonocítica/metabolismo , Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Feminino , Humanos , Índia , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/fisiologia , Leishmaniose Cutânea/parasitologia , Lipocalina-2/genética , Lipocalina-2/metabolismo , Macrófagos/metabolismo , Masculino , Monócitos/metabolismo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores da Transferrina/genética , Receptores da Transferrina/metabolismo , Adulto Jovem
14.
Parasitol Res ; 119(2): 649-657, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31897791

RESUMO

Leishmania donovani is the causative agent of visceral leishmaniasis. Annually, 500 million new cases of infection are reported mainly in poor communities, decreasing the interest of the pharmaceutical industries. Therefore, the repositioning of new drugs is an ideal strategy to fight against these parasites. SQ109, a compound in phase IIb/III of clinical trials to treat resistant Mycobacterium tuberculosis, has a potent effect against Trypanosoma cruzi, responsible for Chagas' disease, and on Leishmania mexicana, the causative agent of cutaneous and muco-cutaneous leishmaniasis. In the latter, the toxic dose against intramacrophagic amastigotes is very low (IC50 ~ 11 nM). The proposed mechanism of action on L. mexicana involves the disruption of the parasite intracellular Ca2+ homeostasis through the collapse of the mitochondrial electrochemical potential (ΔΨm). In the present work, we show a potent effect of SQ109 on L. donovani, the parasite responsible for visceral leishmaniasis, the more severe and uniquely lethal form of these infections, obtaining a toxic effect on amastigotes inside macrophages even lower to that obtained in L. mexicana (IC50 of 7.17 ± 0.09 nM) and with a selectivity index > 800, even higher than in L. mexicana. We also demonstrated for first time that SQ109, besides collapsing ΔΨm of the parasite, induced a very rapid damage to the parasite acidocalcisomes, essential organelles involved in the bioenergetics and many other important functions, including Ca2+ homeostasis. Both effects of the drug on these organelles generated a dramatic increase in the intracellular Ca2+ concentration, causing parasite death.


Assuntos
Adamantano/análogos & derivados , Etilenodiaminas/farmacologia , Leishmania donovani/efeitos dos fármacos , Leishmaniose Cutânea/tratamento farmacológico , Leishmaniose Visceral/tratamento farmacológico , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Adamantano/farmacologia , Animais , Proliferação de Células , Doença de Chagas/tratamento farmacológico , Citoplasma , Humanos , Leishmania mexicana/efeitos dos fármacos , Leishmaniose Cutânea/parasitologia , Leishmaniose Visceral/parasitologia , Macrófagos/parasitologia , Mitocôndrias , Trypanosoma cruzi/efeitos dos fármacos
15.
Molecules ; 25(3)2020 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-31979089

RESUMO

A chemically diverse range of novel tetraoxanes was synthesized and evaluated in vitro against intramacrophage amastigote forms of Leishmania donovani. All 15 tested tetraoxanes displayed activity, with IC50 values ranging from 2 to 45 µm. The most active tetraoxane, compound LC140, exhibited an IC50 value of 2.52 ± 0.65 µm on L. donovani intramacrophage amastigotes, with a selectivity index of 13.5. This compound reduced the liver parasite burden of L. donovani-infected mice by 37% after an intraperitoneal treatment at 10 mg/kg/day for five consecutive days, whereas miltefosine, an antileishmanial drug in use, reduced it by 66%. These results provide a relevant basis for the development of further tetraoxanes as effective, safe, and cheap drugs against leishmaniasis.


Assuntos
Antiprotozoários/química , Antiprotozoários/uso terapêutico , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/patogenicidade , Tetraoxanos/química , Tetraoxanos/uso terapêutico , Animais , Leishmaniose/tratamento farmacológico , Leishmaniose/parasitologia , Camundongos , Fosforilcolina/análogos & derivados , Fosforilcolina/química , Fosforilcolina/uso terapêutico
16.
J Enzyme Inhib Med Chem ; 35(1): 432-459, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31899980

RESUMO

A series of new 2,4-bis[(substituted-aminomethyl)phenyl]quinoline, 1,3-bis[(substituted-aminomethyl)phenyl]isoquinoline, and 2,4-bis[(substituted-aminomethyl)phenyl]quinazoline derivatives was designed, synthesised, and evaluated in vitro against three protozoan parasites (Plasmodium falciparum, Leishmania donovani, and Trypanosoma brucei brucei). Biological results showed antiprotozoal activity with IC50 values in the µM range. In addition, the in vitro cytotoxicity of these original molecules was assessed with human HepG2 cells. The quinoline 1c was identified as the most potent antimalarial candidate with a ratio of cytotoxic to antiparasitic activities of 97 against the P. falciparum CQ-sensitive strain 3D7. The quinazoline 3h was also identified as the most potent trypanosomal candidate with a selectivity index (SI) of 43 on T. brucei brucei strain. Moreover, as the telomeres of the parasites P. falciparum and Trypanosoma are possible targets of this kind of nitrogen heterocyclic compounds, we have also investigated stabilisation of the Plasmodium and Trypanosoma telomeric G-quadruplexes by our best compounds through FRET melting assays.


Assuntos
Antiprotozoários/química , Antiprotozoários/farmacologia , Desenho de Fármacos , Quinolinas/química , Quinolinas/farmacologia , Antiprotozoários/síntese química , Células Hep G2 , Humanos , Leishmania donovani/efeitos dos fármacos , Plasmodium falciparum/efeitos dos fármacos , Quinolinas/síntese química , Relação Estrutura-Atividade , Trypanosoma brucei brucei/efeitos dos fármacos
17.
J Med Chem ; 63(2): 756-783, 2020 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-31846577

RESUMO

From a high-throughput screen of 42 444 known human kinases inhibitors, a pyrazolo[1,5-b]pyridazine scaffold was identified to begin optimization for the treatment of human African trypanosomiasis. Previously reported data for analogous compounds against human kinases GSK-3ß, CDK-2, and CDK-4 were leveraged to try to improve the selectivity of the series, resulting in 23a which showed selectivity for T. b. brucei over these three human enzymes. In parallel, properties known to influence the absorption, distribution, metabolism, and excretion (ADME) profile of the series were optimized resulting in 20g being progressed into an efficacy study in mice. Though 20g showed toxicity in mice, it also demonstrated CNS penetration in a PK study and significant reduction of parasitemia in four out of the six mice.


Assuntos
Piridazinas/síntese química , Piridazinas/farmacologia , Tripanossomicidas/síntese química , Tripanossomicidas/farmacologia , Tripanossomíase Africana/tratamento farmacológico , Animais , Sobrevivência Celular/efeitos dos fármacos , Cristalografia por Raios X , Quinase 2 Dependente de Ciclina/antagonistas & inibidores , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Reposicionamento de Medicamentos , Glicogênio Sintase Quinase 3 beta/antagonistas & inibidores , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Ensaios de Triagem em Larga Escala , Humanos , Leishmania donovani/efeitos dos fármacos , Camundongos , Modelos Moleculares , Piridazinas/farmacocinética , Ratos , Relação Estrutura-Atividade , Especificidade por Substrato , Distribuição Tecidual , Tripanossomicidas/farmacocinética , Trypanosoma brucei brucei/efeitos dos fármacos , Tripanossomíase Africana/parasitologia
18.
Fitoterapia ; 140: 104420, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31733344

RESUMO

We report for the first time the isolation of 2-furyl(phenyl)methanol (5) from the chloroform extracts of the Atractylis gummifera roots. A. gummifera is a thistle belonging to the Asteraceae family that produces the ent-kaurane diterpenoid glycoside atractyloside (ATR). ATR (1) was isolated and chemically modified to obtain its aglycone atractyligenin (2) and the methylated derivatives ATR-OMe (3) and genine-OMe (4). The compounds 1-5 were structurally characterised and evaluated against the intracellular amastigote, cultured within macrophages, and the extracellular promastigote of Leishmania donovani, the protozoan parasite responsible for the highly infective disease visceral leishmaniasis, which is fatal if untreated. The 2-furyl(phenyl)methanol 5 exhibited notable activity against the promastigote.


Assuntos
Antiprotozoários/farmacologia , Atractylis/química , Leishmania donovani/efeitos dos fármacos , Metanol/farmacologia , Animais , Antiprotozoários/isolamento & purificação , Itália , Macrófagos/parasitologia , Metanol/análogos & derivados , Metanol/isolamento & purificação , Camundongos Endogâmicos BALB C , Estrutura Molecular , Compostos Fitoquímicos/isolamento & purificação , Compostos Fitoquímicos/farmacologia , Extratos Vegetais , Rizoma/química
19.
Eur J Med Chem ; 186: 111860, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31759728

RESUMO

Methionine aminopeptidase 1 of Leishmania donovani (LdMetAP1) is a novel antileishmanial target for its role in vital N-terminal methionine processing. After LdMetAP1 expression and purification, we employed a series of biochemical assays to determine optimal conditions for catalysis, metal dependence and substrate preferences for this ubiquitous enzyme. Screening of newly synthesized quinoline-carbaldehyde derivatives in inhibition assays led to the identification of HQ14 and HQ15 as novel and specific inhibitors for LdMetAP1 which compete with substrate for binding to the catalytic active site. Both leads bind LdMetAP1 with high affinity and possess druglikeness. Biochemical studies suggested HQ14 and HQ15 to be comparatively less effective against purified HsMetAP1 and showed no or less toxicity. We further show selectivity and inhibition of lead inhibitors is sensed through a non-catalytic Thr residue unique to LdMetAP1. Finally, structural studies highlight key differences in the binding modes of HQ14 and HQ15 to LdMetAP1 and HsMetAP1 providing structural basis for differences in inhibition. The study demonstrates the feasibility of deploying small drug like molecules to selectively target the catalytic activity of LdMetAP1 which may provide an effective treatment of leishmaniasis.


Assuntos
Aldeídos/farmacologia , Aminopeptidases/antagonistas & inibidores , Antiprotozoários/farmacologia , Leishmania donovani/efeitos dos fármacos , Quinolinas/farmacologia , Aldeídos/síntese química , Aldeídos/química , Aminopeptidases/metabolismo , Antiprotozoários/síntese química , Antiprotozoários/química , Relação Dose-Resposta a Droga , Leishmania donovani/enzimologia , Estrutura Molecular , Testes de Sensibilidade Parasitária , Quinolinas/síntese química , Quinolinas/química , Relação Estrutura-Atividade
20.
J Ethnopharmacol ; 247: 112270, 2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-31589965

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Primates forage on a variety of plant parts to balance their dietary intake to meet requirements of energy, nutrition and maintenance, however the reason(s) leading them to ingest some plants which have no nutritional value and/or contain bioactive or even toxic secondary metabolites is recently gaining closer attention. The growing literature suggests that primates consume plants for medicinal purposes (self-medication) as well, particularly when infected with parasites and pathogens (bacteria, viruses, microbes). Interestingly, some of the plants they consume are also used by humans for similar purposes or may have potential uses for humans. MATERIALS AND METHODS: As part of a 16-month study of the parasite ecology of a sub-species of Japanese macaques (Macaca fuscata yakui) on the island of Yakushima, we surveyed their feeding habits and collected a subset of plants and plant parts observed being ingested by macaques. The ethnomedicinal value of these plants was surveyed and methanolic extracts of 45 plant parts were tested in vitro against important parasites of humans, including four protozoan parasites Plasmodium falciparum, Trypanosoma brucei rhodesiense, T. cruzi and Leishmania donovani, and the trematode flatworm Schistosoma mansoni. Potential toxicity of the extracts was also assessed on mammalian cells. RESULTS: A wide range of ethnomedicinal uses in Asia for these plants is noted, with 37% associated with the treatment of parasites, pathogens and related symptoms. Additionally, the 45 extracts tested showed broad and significant activity against our test organisms. All extracts were active against T. b. rhodesiense. The majority (over 80%) inhibited the growth of P. falciparum and L. donovani. Half of the extracts also displayed antiprotozoal potential against T. cruzi while only several extracts were active against both larval and adult stages of S. mansoni. Cytotoxicity was generally low, although several extracts lacked specific toxicity to test parasites. CONCLUSIONS: Our results indicated a number of plants and their parts to have antiparasitic activity not previously reported in the ethnopharmacological literature. Enhanced understanding of the primate diets, particularly during periods of intensified parasite infection risk may help to further narrow down plants of interest for lead compound development. The study of animal self-medication is a complementary approach, with precedence, to drug discovery of new lead drug compounds against human parasitic diseases.


Assuntos
Anti-Helmínticos/farmacologia , Antiprotozoários/farmacologia , Macaca fuscata/parasitologia , Extratos Vegetais/farmacologia , Plantas Medicinais/química , Infecções Protozoárias em Animais/tratamento farmacológico , Esquistossomose mansoni/tratamento farmacológico , Animais , Anti-Helmínticos/uso terapêutico , Antiprotozoários/uso terapêutico , Etnofarmacologia , Comportamento Alimentar , Feminino , Ilhas , Japão , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/isolamento & purificação , Masculino , Medicina Tradicional/métodos , Testes de Sensibilidade Parasitária , Extratos Vegetais/uso terapêutico , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/isolamento & purificação , Infecções Protozoárias em Animais/parasitologia , Schistosoma mansoni/isolamento & purificação , Esquistossomose mansoni/parasitologia , Esquistossomose mansoni/veterinária , Automedicação/veterinária , Testes de Toxicidade , Trypanosoma brucei rhodesiense/isolamento & purificação , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/isolamento & purificação
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