Mechanisms of Arsenical and Diamidine Uptake and Resistance in Trypanosoma brucei

Abstract

Sleeping sickness, caused by Trypanosoma brucei spp., has become resurgent in sub-Saharan Africa. Moreover, there is an alarming increase in treatment failures with melarsoprol, the principal agent used against late-stage sleeping sickness. In T. brucei, the uptake of melarsoprol as well as diamidines is thought to be mediated by the P2 aminopurine transporter, and loss of P2 function has been implicated in resistance to these agents. The trypanosomal gene TbAT1 has been found to encode a P2-type transporter when expressed in yeast. Here we investigate the role of TbAT1 in drug uptake and drug resistance in T. brucei by genetic knockout of TbAT1. Tbat1-null trypanosomes were deficient in P2-type adenosine transport and lacked adenosine-sensitive transport of pentamidine and melaminophenyl arsenicals. However, the null mutants were only slightly resistant to melaminophenyl arsenicals and pentamidine, while resistance to other diamidines such as diminazene was more pronounced. Nevertheless, the reduction in drug sensitivity might be of clinical significance, since mice infected with tbat1-null trypanosomes could not be cured with 2 mg of melarsoprol/kg of body weight for four consecutive days, whereas mice infected with the parental line were all cured by using this protocol. Two additional pentamidine transporters, HAPT1 and LAPT1, were still present in the null mutant, and evidence is presented that HAPT1 may be responsible for the residual uptake of melaminophenyl arsenicals. High-level arsenical resistance therefore appears to involve the loss of more than one transporter.

Aquaglyceroporin 2 controls susceptibility to melarsoprol and pentamidine in African trypanosomes

“…AT1 has been found to be mutated, deleted, or down-regulated in melarsoprol-resistant trypanosomes (10,42) but AT1 knockout generated cells with only a twofold increased resistance to both melarsoprol and pentamidine (14), and there is evidence for resistance that is independent of AT1 disruption (43,44). Our results now show that AQP2, when defective, could explain cases of innate MPXR and/or acquired MPXR of clinical relevance.…”

“…The same P2/AT1 mediates pentamidine transport in trypanosomes (11,12) and when expressed in S. cerevisiae (13). AT1 gene knockout generated cells with approximately twofold increased resistance to both melarsoprol and pentamidine (14) and AT1 knockdown, using RNA interference (RNAi), increased resistance to melarsoprol (5, 15) but has not been shown to significantly increase resistance to pentamidine. Additional T. brucei transporters, NT11.1 and NT12.1, can transport pentamidine when expressed in a heterologous system (16) and high and low-affinity pentamidine transporters (HAPT1 and LAPT1, respectively) have been described (12,17).…”

The adaptive potential of a survival artist: characterization of thein vitrointeractions ofToxoplasma gondiitachyzoites with di-cationic compounds in human fibroblast cell cultures

“…A prominent ABC transporter, Pglycoprotein (TgABC.B1 and TgABC.B2) potentially mediating transport of the drug out of the cell, are associated with the membrane in tachyzoites, and is constitutively expressed in all 3 virulent types. On the other hand, another adaptation mechanism could involve the reduced uptake of drug, due to the downregulation of certain adenosine transporter activities, which are involved in diamidine uptake (Matovu et al 2003;Witola et al 2004). For instance, homologues to the P2 aminopurine transporter in T. brucei are found in the Toxoplasma genome (www.toxodb.…”

Molecular tools for the rapid detection of drug resistance in animal trypanosomes

“…The role of the P2-type purine transporter in the uptake of arsenical diamidines, pentamidine and DA by T. brucei, T. evansi and T. equiperdum, and the consequences of inhibition, knocking down or silencing this gene have been extensively described and reviewed in the literature [33], [34], [35], [36], [37], [38], [39], [40], [41], [42] and [43]. In addition to this resistance mechanism, a novel gene, TeDR40, might be a factor contributing to high DA-resistance in T. evansi [44].…”