RESUMEN
Human African trypanosomiasis or sleeping sickness is a deadly disease endemic in sub-Saharan Africa, caused by single-celled protozoan parasites. Although it has been targeted for elimination by 2020, this will only be realized if diagnosis can be improved to enable identification and treatment of afflicted patients. Existing techniques of detection are restricted by their limited field-applicability, sensitivity and capacity for automation. Microfluidic-based technologies offer the potential for highly sensitive automated devices that could achieve detection at the lowest levels of parasitemia and consequently help in the elimination programme. In this work we implement an electrokinetic technique for the separation of trypanosomes from both mouse and human blood. This technique utilises differences in polarisability between the blood cells and trypanosomes to achieve separation through opposed bi-directional movement (cell counterflow). We combine this enrichment technique with an automated image analysis detection algorithm, negating the need for a human operator.
Asunto(s)
Trypanosoma/aislamiento & purificación , Tripanosomiasis Africana/diagnóstico , Algoritmos , Animales , Electroforesis/métodos , Humanos , Ratones , Parasitemia , Tripanosomiasis Africana/sangre , Tripanosomiasis Africana/parasitologíaRESUMEN
Acivicin analogues with an increased affinity for CTP synthetase (CTPS) were designed as potential new trypanocidal agents. The inhibitory activity against CTPS can be improved by increasing molecular complexity, by inserting groups able to establish additional interactions with the binding pocket of the enzyme. This strategy has been pursued with the synthesis of α-amino-substituted analogues of Acivicin and N1-substituted pyrazoline derivatives. In general, there is direct correlation between the enzymatic activity and the in vitro anti-trypanosomal efficacy of the derivatives studied here. However, this cannot be taken as a general rule, as other important factors may play a role, notably the ability of uptake/diffusion of the molecules into the trypanosomes.
Asunto(s)
Ligasas de Carbono-Nitrógeno/antagonistas & inhibidores , Isoxazoles/química , Isoxazoles/farmacología , Tripanocidas/química , Tripanocidas/farmacología , Trypanosoma brucei brucei/efectos de los fármacos , Ligasas de Carbono-Nitrógeno/metabolismo , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Células HeLa , Humanos , Simulación del Acoplamiento Molecular , Pirazoles/química , Pirazoles/farmacología , Trypanosoma brucei brucei/enzimología , Tripanosomiasis Africana/tratamiento farmacológicoRESUMEN
The first convenient synthesis of enantiomerically pure (αS,5S)-α-amino-3-bromo-4,5-dihydroisoxazol-5-yl acetic acid (3-bromoacivicin) is described. We demonstrate that 3-bromoacivicin is a CTP synthetase inhibitor three times as potent as its 3-chloro analogue, the natural antibiotic acivicin. Because CTP synthetase was suggested to be a potential drug target in African trypanosomes, the inâ vitro/inâ vivo antitrypanosomal activity of 3-bromoacivicin was assessed in comparison with acivicin. Beyond expectation, we observed a 12-fold enhancement in the inâ vitro antitrypanosomal activity, while toxicity against mammalian cells remained unaffected. Despite its good inâ vitro activity and selectivity, 3-bromoacivicin proved to be trypanostatic and failed to completely eradicate the infection when tested inâ vivo at its maximum tolerable dose.
Asunto(s)
Ligasas de Carbono-Nitrógeno/antagonistas & inhibidores , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Isoxazoles/síntesis química , Isoxazoles/farmacología , Evaluación Preclínica de Medicamentos , Técnicas In VitroRESUMEN
A series of unsaturated Mannich bases possessing two electrophilic sites was recently identified as irreversible inhibitors of trypanothione reductase from Trypanosoma cruzi. New derivatives were synthesized by modifying the substitution pattern on the aromatic ring and by incorporating the melamine motif of melarsoprol. Their affinity to P2 transporter and their trypanocidal properties have been studied using three strains expressing various purine transporters. While the melamine derivatives showed some affinity to the P2 transporter, unsaturated Mannich bases without the melamine motif showed excellent potencies against pentamidine-resistant strains of T. brucei brucei suggesting alternative drug uptake routes. The Michael acceptor properties of the three most active compounds towards glutathione correlated with the observed trypanocidal activities.