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A new approach to chemotherapy: drug-induced differentiation kills African trypanosomes.

ABSTRACT: Human African trypanosomiasis (sleeping sickness) is a neglected tropical disease caused by Trypanosoma brucei spp. The parasites are transmitted by tsetse flies and adapt to their different hosts and environments by undergoing a series of developmental changes. During differentiation, the trypanosome alters its protein coat. Bloodstream form trypanosomes in humans have a coat of variant surface glycoprotein (VSG) that shields them from the immune system. The procyclic form, the first life-cycle stage to develop in the tsetse fly, replaces the VSG coat by procyclins; these proteins do not protect the parasite from lysis by serum components. Our study exploits the parasite-specific process of differentiation from bloodstream to procyclic forms to screen for potential drug candidates. Using transgenic trypanosomes with a reporter gene in a procyclin locus, we established a whole-cell assay for differentiation in a medium-throughput format. We screened 7,495 drug-like compounds and identified 28 hits that induced expression of the reporter and loss of VSG at concentrations in the low micromolar range. Small molecules that induce differentiation to procyclic forms could facilitate studies on the regulation of differentiation as well as serving as scaffolds for medicinal chemistry for new treatments for sleeping sickness.

SUBMITTER: Wenzler T

PROVIDER: S-EPMC4773815 | biostudies-other | 2016 Mar

REPOSITORIES: biostudies-other

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A new approach to chemotherapy: drug-induced differentiation kills African trypanosomes.

Wenzler Tanja T Schumann Burkard Gabriela G Schmidt Remo S RS Mäser Pascal P Bergner Andreas A Roditi Isabel I Brun Reto R

Scientific reports 20160302

Human African trypanosomiasis (sleeping sickness) is a neglected tropical disease caused by Trypanosoma brucei spp. The parasites are transmitted by tsetse flies and adapt to their different hosts and environments by undergoing a series of developmental changes. During differentiation, the trypanosome alters its protein coat. Bloodstream form trypanosomes in humans have a coat of variant surface glycoprotein (VSG) that shields them from the immune system. The procyclic form, the first life-cycle ...[more]

PMID: 26931380

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Project description:The African trypanosome has evolved mechanisms to adapt to changes in nutrient availability that occur during its life cycle. During transition from mammalian blood to insect vector gut, parasites experience a rapid reduction in environmental glucose. Here we describe how pleomorphic parasites respond to glucose depletion with a focus on parasite changes in energy metabolism and growth. Long slender bloodstream form parasites were rapidly killed as glucose concentrations fell, while short stumpy bloodstream form parasites persisted to differentiate into the insect-stage procyclic form parasite. The rate of differentiation was lower than that triggered by other cues but reached physiological rates when combined with cold shock. Both differentiation and growth of resulting procyclic form parasites were inhibited by glucose and nonmetabolizable glucose analogs, and these parasites were found to have upregulated amino acid metabolic pathway component gene expression. In summary, glucose transitions from the primary metabolite of the blood-stage infection to a negative regulator of cell development and growth in the insect vector, suggesting that the hexose is not only a key metabolic agent but also an important signaling molecule.IMPORTANCE As the African trypanosome Trypanosoma brucei completes its life cycle, it encounters many different environments. Adaptation to these environments includes modulation of metabolic pathways to parallel the availability of nutrients. Here, we describe how the blood-dwelling life cycle stages of the African trypanosome, which consume glucose to meet their nutritional needs, respond differently to culture in the near absence of glucose. The proliferative long slender parasites rapidly die, while the nondividing short stumpy parasite remains viable and undergoes differentiation to the next life cycle stage, the procyclic form parasite. Interestingly, a sugar analog that cannot be used as an energy source inhibited the process. Furthermore, the growth of procyclic form parasite that resulted from the event was inhibited by glucose, a behavior that is similar to that of parasites isolated from tsetse flies. Our findings suggest that glucose sensing serves as an important modulator of nutrient adaptation in the parasite.

Project description:Bloodstream forms of four populations of the livestock pathogen Trypanosoma congolense, isolated from different natural infections, have been shown to exhibit a wide range of sensitivities to the trypanocide isometamidium chloride (Samorin(R)). In mice the 50% curative doses (CD50) for Samorin range from 0.007 to 20 mg/kg body weight. Uptake of isometamidium chloride demonstrated Michaelis-Menten-type kinetics in all the populations, with Km values in the range 0.35-0.87 microM, and Vmax varied from 17 to 216 pmol/min per 10(8) cells. The magnitude of Vmax was correlated with sensitivity to the drug. In contrast, no correlation was observed between Km values and drug sensitivity. Pulse-chase experiments indicated two compartments for accumulation of drug. The first consists of freely diffusible drug that is invariant between populations; the other consists of retained isometamidium, which is of variable magnitude between the populations and is correlated with drug sensitivity. Autoradiography and fluorescence microscopy demonstrated initial, rapid accumulation of the drug within the mitochondrion, specifically the kinetoplast. In a drug-sensitive population of T. congolense, agents affecting mitochondrial function were shown to produce dose-dependent inhibition of mitochondrial membrane potential (DeltaPsimito), as measured by the accumulation of the lipophilic cations [3H]methyltriphenylphosphonium iodide or rhodamine 123. The agents also produced parallel inhibition of isometamidium uptake, suggesting an involvement of DeltaPsimito in the accumulation of the drug. When characterized in each of the four populations, the spontaneous DeltaPsimito was shown to be characteristic of each population and was correlated with Vmax for drug uptake and sensitivity to the drug in vitro and in vivo. We therefore conclude that in T. congolense DeltaPsimito is an important determinant of the rate and accumulation of the trypanocide isometamidium chloride. Populations of this trypanosome species vary with respect to DeltaPsimito, which is correlated with sensitivity to isometamidium. We suggest that when exposed to drug, the selection of such populations represents a novel mechanism of drug resistance in protozoan parasites.

Dataset Information

A new approach to chemotherapy: drug-induced differentiation kills African trypanosomes.

Publications

A new approach to chemotherapy: drug-induced differentiation kills African trypanosomes.

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