Cloning and Characterization of Trypanosoma congolense and T. vivax Nucleoside Transporters Reveal the Potential of P1-Type Carriers for the Discovery of Broad-Spectrum Nucleoside-Based Therapeutics against Animal African Trypanosomiasis

Abstract

African Animal Trypanosomiasis (AAT), caused predominantly by Trypanosoma brucei brucei, T. vivax and T. congolense, is a fatal livestock disease throughout Sub-Saharan Africa. Treatment options are very limited and threatened by resistance. Tubercidin (7-deazaadenosine) analogs have shown activity against individual parasites but viable chemotherapy must be active against all three species. Divergence in sensitivity to nucleoside antimetabolites could be caused by differences in nucleoside transporters. Having previously characterized the T. brucei nucleoside carriers, we here report the functional expression and characterization of the main adenosine transporters of T. vivax (TvxNT3) and T. congolense (TcoAT1/NT10), in a Leishmania mexicana cell line (‘SUPKO’) lacking adenosine uptake. Both carriers were similar to the T. brucei P1-type transporters and bind adenosine mostly through interactions with N3, N7 and 3′-OH. Expression of TvxNT3 and TcoAT1 sensitized SUPKO cells to various 7-substituted tubercidins and other nucleoside analogs although tubercidin itself is a poor substrate for P1-type transporters. Individual nucleoside EC50s were similar for T. b. brucei, T. congolense, T. evansi and T. equiperdum but correlated less well with T. vivax. However, multiple nucleosides including 7-halogentubercidines displayed pEC50>7 for all species and, based on transporter and anti-parasite SAR analyses, we conclude that nucleoside chemotherapy for AAT is viable.

In Vitro and In Vivo Trypanocidal Efficacy of Nitrofuryl- and Nitrothienylazines

“…Because the nitrofuranylazines showed the best trypanocidal activity against the animal trypanosome T. congolense IL3000 (Figure 2), structure−activity relationships (SARs) were observed relative to the said trypanosome. Differences in drug sensitivity may result from variances in the parasites' capacity to internalize, 59 activate, 60 and/or metabolize 61 drug candidates. Moreover, based on the promising in vitro trypanocidal activity and selectivity of derivatives 4a, 7a, and 8b (Table 2), these azines were selected for in vivo treatment efficacy assessment in T. congolense IL3000 infected mice.…”

Targeting the nucleotide metabolism of Trypanosoma brucei and other trypanosomatids

“…The recognition patterns of P1 and P2 will be further described in the drug development sections. Interestingly, T. congolense and T. vivax that cause animal trypanosomiasis lack P2 altogether and have only one major P1 transporting activity in each species (Ungogo et al 2023 ). The single T. congolense P1 transporter is called Tco AT1 or Tco NT10 (not to be confused with Tb AT1 from T. brucei , which is a P2 transporter), and the only T. vivax P1 transporter that could be confirmed to be active is called Tvx NT3.…”

“…Transport studies showed that both Tco AT1 and Tvx NT3 are purine-specific and recognize typical P1 substrates (adenosine, inosine, and guanosine), but Tvx NT3 has a broader substrate specificity and can also bind purine nucleobases (not tested as substrates). Similarly to the T. brucei P1 transporters (but unlike P2), Tco AT1 and Tvx NT3 does not seem to transport drugs tested from the diamidine and melanophenyl arsenical families (Munday et al 2013 , Ungogo et al 2023 ). In addition to the purine nucleoside transporters, all three species have several genes encoding purine nucleobase transporters, although they have only been characterized in T. brucei .…”

Overcoming Biological Barriers: Importance of Membrane Transporters in Homeostasis, Disease and Disease Treatment

“…This system provides an ideal null background for the expression and characterization of single equilibrative nucleoside transporter (ENT) genes. Using the same null background Leishmania mexicana cell line, in the manuscript “Cloning and Characterization of Trypanosoma congolense and T. vivax Nucleoside Transporters Reveal the Potential of P1-Type Carriers for the Discovery of Broad-Spectrum Nucleoside-Based Therapeutics against Animal African Trypanosomiasis” Ungogo et al [ 7 ] investigated the druggability of the main adenosine transporters (purine nucleoside transporter NT3) of T. vivax (TvxNT3) and T. congolense (TcoAT1/NT10) by 7-substituted tubercidins and other nucleoside analogs. They showed that chemotherapy for Animal African Trypanosomiasis is possible.…”