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1.
Front Immunol ; 11: 1113, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32582193

RESUMO

Type I interferons (IFNs) induced by an endogenous Leishmania RNA virus or exogenous viral infections have been shown to exacerbate infections with New World Cutaneous Leishmania parasites, however, the impact of type I IFNs in visceral Leishmania infections and implicated mechanisms remain to be unraveled. This study assessed the impact of type I IFN on macrophage infection with L. infantum and L. donovani and the implication of sialoadhesin (Siglec-1/CD169, Sn) as an IFN-inducible surface receptor. Stimulation of bone marrow-derived macrophages with type I IFN (IFN-α) significantly enhanced susceptibility to infection of reference laboratory strains and a set of recent clinical isolates. IFN-α particularly enhanced promastigote uptake. Enhanced macrophage susceptibility was linked to upregulated Sn surface expression as a major contributing factor to the infection exacerbating effect of IFN-α. Stimulation experiments in Sn-deficient macrophages, macrophage pretreatment with a monoclonal anti-Sn antibody or a novel bivalent anti-Sn nanobody and blocking of parasites with soluble Sn restored normal susceptibility levels. Infection of Sn-deficient mice with bioluminescent L. infantum promastigotes revealed a moderate, strain-dependent role for Sn during visceral infection under the used experimental conditions. These data indicate that IFN-responsive Sn expression can enhance the susceptibility of macrophages to infection with visceral Leishmania promastigotes and that targeting of Sn may have some protective effects in early infection.

2.
ACS Infect Dis ; 6(8): 2045-2056, 2020 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-32568511

RESUMO

Human African trypanosomiasis is a neglected tropical disease caused by Trypanosoma brucei parasites. These protists are unable to produce the purine ring, making them vulnerable to the effects of purine nucleoside analogues. Starting from 3'-deoxytubercidin (5), a lead compound with activity against central-nervous-stage human African trypanosomiasis, we investigate the structure-activity relationships of the purine and ribofuranose rings. The purine ring tolerated only modifications at C7, while from the many alterations of the 3'-deoxyribofuranosyl moiety only the arabino analogue 48 showed pronounced antitrypanosomal activity. Profiling of the most potent analogues against resistant T. brucei strains (resistant to pentamidine, diminazene, and isometamidium) showed reduced dependence on uptake mediated by the P2 aminopurine transporter relative to 5. The introduction of a 7-substituent confers up to 10-fold increased affinity for the P1 nucleoside transporter while generally retaining high affinity for P2. Four of the most promising analogues were found to be metabolically stable, earmarking them as suitable backup analogues for lead 5.

3.
Curr Opin Immunol ; 66: 65-73, 2020 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-32446136

RESUMO

Human African trypanosomes rely for their transmission on tsetse flies (Glossina sp.) that inoculate parasites into the skin during blood feeding. The absence of a protective vaccine, limited knowledge about the infection immunology, and the existence of asymptomatic carriers sustaining transmission are major outstanding challenges towards elimination. All these relate to the skin where (i) parasites persist and transmit to tsetse flies and (ii) a successful vaccination strategy should ideally be effective. Host immune processes and parasite strategies that underlie early infection and skin tropism are essential aspects to comprehend the transmission-success of trypanosomes and the failure in vaccine development. Recent insights into the early infection establishment may pave the way to novel strategies aimed at blocking transmission.

4.
Eur J Med Chem ; 188: 112018, 2020 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-31931339

RESUMO

African trypanosomiasis, a deadly infectious disease caused by the protozoan Trypanosoma brucei spp., is spread to new hosts by bites of infected tsetse flies. Currently approved therapies all have their specific drawbacks, prompting a search for novel therapeutic agents. T. brucei lacks the enzymes necessary to forge the purine ring from amino acid precursors, rendering them dependent on the uptake and interconversion of host purines. This dependency renders analogues of purines and corresponding nucleosides an interesting source of potential anti-T. brucei agents. In this study, we synthesized and evaluated a series of 7-substituted 7-deazainosine derivatives and found that 6-O-alkylated analogues in particular showed highly promising in vitro activity with EC50 values in the mid-nanomolar range. SAR investigation of the O-alkyl chain showed that antitrypanosomal activity increased, and also cytotoxicity, with alkyl chain length, at least in the linear alkyl chain series. However, this could be attenuated by introducing a terminal branch point, resulting in the highly potent and selective analogues, 36, 37 and 38. No resistance related to transporter-mediated uptake could be identified, earmarking several of these analogues for further in vivo follow-up studies.


Assuntos
Descoberta de Drogas , Inosina/análogos & derivados , Nucleosídeos/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma brucei brucei/efeitos dos fármacos , Alquilação , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Inosina/síntese química , Inosina/química , Inosina/farmacologia , Estrutura Molecular , Nucleosídeos/síntese química , Nucleosídeos/química , Testes de Sensibilidade Parasitária , Relação Estrutura-Atividade , Tripanossomicidas/síntese química , Tripanossomicidas/química
5.
Nat Commun ; 10(1): 5564, 2019 12 05.
Artigo em Inglês | MEDLINE | ID: mdl-31804484

RESUMO

African trypanosomiasis is a disease caused by Trypanosoma brucei parasites with limited treatment options. Trypanosoma is unable to synthesize purines de novo and relies solely on their uptake and interconversion from the host, constituting purine nucleoside analogues a potential source of antitrypanosomal agents. Here we combine structural elements from known trypanocidal nucleoside analogues to develop a series of 3'-deoxy-7-deazaadenosine nucleosides, and investigate their effects against African trypanosomes. 3'-Deoxytubercidin is a highly potent trypanocide in vitro and displays curative activity in animal models of acute and CNS-stage disease, even at low doses and oral administration. Whole-genome RNAi screening reveals that the P2 nucleoside transporter and adenosine kinase are involved in the uptake and activation, respectively, of this analogue. This is confirmed by P1 and P2 transporter assays and nucleotide pool analysis. 3'-Deoxytubercidin is a promising lead to treat late-stage sleeping sickness.


Assuntos
Desoxiadenosinas/farmacologia , Proteínas de Transporte de Nucleosídeos/metabolismo , Proteínas de Protozoários/metabolismo , Trypanosoma brucei brucei/efeitos dos fármacos , Tripanossomíase Africana/tratamento farmacológico , Tubercidina/farmacologia , Animais , Antimetabólitos Antineoplásicos/química , Antimetabólitos Antineoplásicos/farmacologia , Linhagem Celular , Sobrevivência Celular , Desoxiadenosinas/química , Quimioterapia Combinada , Feminino , Humanos , Camundongos , Estrutura Molecular , Proteínas de Transporte de Nucleosídeos/genética , Proteínas de Protozoários/genética , Tripanossomicidas/química , Tripanossomicidas/farmacologia , Trypanosoma brucei brucei/genética , Trypanosoma brucei brucei/fisiologia , Tripanossomíase Africana/parasitologia , Tubercidina/química
6.
Sci Rep ; 8(1): 11203, 2018 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-30046157

RESUMO

In this study, Trypanosoma brucei was naturally transmitted to mice through the bites of infected Glossina morsitans tsetse flies. Neutrophils were recruited rapidly to the bite site, whereas monocytes were attracted more gradually. Expression of inflammatory cytokines (il1b, il6), il10 and neutrophil chemokines (cxcl1, cxcl5) was transiently up-regulated at the site of parasite inoculation. Then, a second influx of neutrophils occurred that coincided with the previously described parasite retention and expansion in the ear dermis. Congenital and experimental neutropenia models, combined with bioluminescent imaging, indicate that neutrophils do not significantly contribute to dermal parasite control and elicit higher systemic parasitemia levels during the infection onset. Engulfment of parasites by neutrophils in the skin was rarely observed and was restricted to parasites with reduced motility/viability, whereas live parasites escaped phagocytosis. To our knowledge, this study represents the first description of a trypanosome infection promoting role of early innate immunological reactions following an infective tsetse fly bite. Our data indicate that the trypanosome is not hindered in its early development and benefits from the host innate responses with the neutrophils being important regulators of the early infection, as already demonstrated for the sand fly transmitted Leishmania parasite.


Assuntos
Derme/parasitologia , Neutrófilos/parasitologia , Trypanosoma brucei brucei/genética , Tripanossomíase Africana/genética , Animais , Quimiocina CXCL1/genética , Quimiocina CXCL5/genética , Derme/metabolismo , Regulação da Expressão Gênica , Mordeduras e Picadas de Insetos/parasitologia , Insetos Vetores/genética , Insetos Vetores/parasitologia , Interleucina-10/genética , Interleucina-1beta/genética , Interleucina-6/genética , Medições Luminescentes , Camundongos , Neutrófilos/metabolismo , Neutrófilos/patologia , Trypanosoma brucei brucei/patogenicidade , Tripanossomíase Africana/parasitologia , Tripanossomíase Africana/transmissão , Moscas Tsé-Tsé/parasitologia , Moscas Tsé-Tsé/patogenicidade
7.
J Mol Diagn ; 20(2): 253-263, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29355825

RESUMO

Several methods have been developed for the detection of Leishmania, mostly targeting the minicircle kinetoplast DNA (kDNA). A new RNA real-time quantitative PCR (qPCR) assay was developed targeting the conserved and highly expressed spliced-leader (SL) mini-exon sequence. This study compared the limits of detection of various real-time PCR assays in hamsters infected with Leishmania infantum, in spiked human blood, and in clinical blood samples from visceral leishmaniasis patients. The SL-RNA assay showed an excellent analytical sensitivity in tissues (0.005 and 0.002 parasites per mg liver and spleen, respectively) and was not prone to false-positive reactions. Evaluation of the SL-RNA assay on clinical samples demonstrated lower threshold cycle values than the kDNA qPCR, an excellent interrun stability of 97%, a 93% agreement with the kDNA assay, and an estimated sensitivity, specificity, and accuracy of 93.2%, 94.3%, and 93.8%, respectively. The SL-RNA qPCR assay was equally efficient for detecting Leishmania major, Leishmania tropica, Leishmania mexicana, Leishmania guayensis, Leishmania panamensis, Leishmania braziliensis, L. infantum, and Leishmania donovani and revealed similar SL-RNA levels in the different species and the occurrence of polycistronic SL-containing transcripts in Viannia species. Collectively, this single SL-RNA qPCR assay enables universal Leishmania detection and represents a particularly useful addition to the widely used kDNA assay in clinical studies in which the detection of viable parasites is pivotal to assess parasitological cure.


Assuntos
DNA de Cinetoplasto/análise , Leishmania infantum/genética , Leishmaniose Visceral/sangue , Leishmaniose Visceral/diagnóstico , Mesocricetus/parasitologia , RNA Líder para Processamento/análise , Reação em Cadeia da Polimerase em Tempo Real/métodos , Animais , Criança , Pré-Escolar , Cricetinae , Confiabilidade dos Dados , Feminino , Humanos , Fígado/parasitologia , Sensibilidade e Especificidade , Baço/parasitologia
8.
PLoS Negl Trop Dis ; 11(5): e0005620, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28505185

RESUMO

BACKGROUND: Since miltefosine monotherapy against visceral leishmaniasis (VL) caused by Leishmania donovani has been discontinued in the Indian subcontinent due to an increase in the number of treatment failures, single dose liposomal amphotericin B is now advocated as a treatment option of choice. Paromomycin-miltefosine combination therapy can be used as substitute first-line treatment in regions without cold-chain potential. Previous laboratory studies in the closely related species Leishmania infantum have demonstrated that paromomycin monotherapy fairly rapidly selects for resistance producing a phenotype with increased fitness. Given the possible clinical implications of these findings for the current field situation, the present study aimed to identify the potential hazards of paromomycin-miltefosine combination therapy. PRINCIPAL FINDINGS: Drug interaction studies using the fixed-ratio isobologram method revealed an indifferent interaction between paromomycin and miltefosine. In hamsters infected with L. infantum, the combination resulted in cumulative efficacy in reducing parasite burdens in the liver, spleen and bone marrow. Selected resistant lines against the single drugs did not display cross-resistance. When the intracellular amastigote stage was repeatedly exposed to the paromomycin-miltefosine combination, either in vitro or in vivo, no significant susceptibility decrease towards either drug was noted. CONCLUSIONS: These results suggest that implementation of paromomycin-miltefosine combination therapy indeed could represent a safe and affordable treatment option for L. donovani VL as miltefosine appears to overrule the anticipated rapid development of PMM resistance.


Assuntos
Antiprotozoários/administração & dosagem , Leishmania infantum/efeitos dos fármacos , Leishmaniose Visceral/tratamento farmacológico , Paromomicina/administração & dosagem , Fosforilcolina/análogos & derivados , Animais , Cricetinae , Resistência a Múltiplos Medicamentos , Quimioterapia Combinada , Feminino , Camundongos , Fosforilcolina/administração & dosagem
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