Investigating Mammalian Tyrosine Phosphatase Inhibitors as Potential ‘Piggyback’ Leads to Target Trypanosoma brucei Transmission

Protein tyrosine phosphatase 1B (PTP1B) is one of the most attractive phosphatase targets in the field of drug discovery. PTP1B is a negative regulator of the insulin and leptin pathways, and mice deficient for this gene show increased sensitivity to insulin and resistance to obesity (1), thus establishing PTP1B as a therapeutic target for the treatment of type 2 diabetes and obesity. Several pharmaceutical companies have been working to develop potent and selective PTP1B inhibitors (2,3) as obesity represents a hugely lucrative market: according to the WHO projections, approximately 2.3 billion adults will be overweight and more than 700 million will be obese by 2015. Trypanosoma brucei protein tyrosine phosphatase 1 (TbPTP1) is 24% identical to human PTP1B at the protein level and possesses 9 of the 10 conserved PTP motifs. TbPTP1 has been identified as a key regulator of the life cycle of African trypanosomes (4), parasites that are the causative agent of Human African trypanosomiasis and of ‘Nagana’, a wasting disease affecting cattle. Both the human and livestock disease generate significant public health and economic burdens throughout 36 sub-Saharan African countries (5). Trypanosoma brucei is transmitted between mammalian hosts by tsetse flies and displays a complex life cycle to survive in these different environmental conditions. In the mammalian bloodstream, ‘stumpy’ form parasites are responsible for disease transmission, as they are the only life cycle form capable of surviving in the vector (6). Once in the tsetse fly midgut, stumpy forms differentiate into procyclic forms, a response that can be reproduced in vitro by a combination of temperature reduction and the addition of citrate/cis-aconitate (7,8). A central component of the signaling pathways contributing to the initiation of differentiation is TbPTP1, which prevents the development from stumpy forms to procyclic forms. A downstream substrate for TbPTP1 is a glycosomal directed phosphatase, TbPIP39, which promotes differentiation when it becomes phosphorylated as a consequence of the inhibition of TbPTP1 (9). Hence, a pharmacological inhibition of TbPTP1 in the mammalian bloodstream would provoke premature differentiation of stumpy forms to procyclic forms, a response that would prove lethal in the parasite. Targeting TbPTP1 would therefore eliminate the population of fly-infective parasites and so prevent disease transmission. Vector control and animal reservoir treatment (10) are strategies currently used to limit the spread of African trypanosomiasis. However, these approaches have limitations that render them unsustainable for the long-term control of disease transmission (11). Consequently, the potential use of a safe and cheap drug to block transmission would be a novel and useful tool for controlling human and animal trypanosomiasis, particularly under epidemic conditions. Moreover, the possibility of ‘piggybacking’ on the extensive efforts in the pharmaceutical industry to develop PTP1B inhibitors would represent a rapid and cost effective route to implementation where preclinical or clinical studies have already been performed (12). This strategy is particularly attractive for sleeping sickness and for other neglected tropical diseases, where economic considerations and the value of potential markets limits investment by the pharmaceutical industry (13). Here, we have tested 19 established PTP1B inhibitors against TbPTP1, including suramin, a front line therapeutic for early stage trypanosomiasis. In each case, their IC50 and Ki values have been calculated, and kinetic parameters determined. Our findings indicate that the PTP1B inhibitors analyzed display similar inhibitory properties against the human and the parasite enzymes, this being consistent with the predicted conservation of their overall 3D structures. Although the low activity of the respective compounds against trypanosomes in culture indicated further refinement is necessary, these analyses validate the general approach of exploiting ‘piggyback’ strategies to control African trypanosomiasis transmission.