Suramin resistance in African Trypanosomes

Drug resistance is a wide-spread phenomenon and affects medical fields from infection biology to oncology. In the protozoan parasite Trypanosoma brucei, the causative agent of sleeping sickness, a number of resistance mechanisms have been described so far, most of them affecting drug uptake. In the presented PhD study I have investigated resistance of trypanosomes against the drug suramin. Suramin is a very old drug but its mode of action in trypanosomes is not well understood. By investigation of suramin-resistant parasites on their genomic and transcriptomic level I aimed to gain new insights into the pathway by which trypanosomes take up suramin and the mode of action of suramin.
I analyzed the transcriptome of a T. brucei rhodesiense line with a very quickly emerging in vitro suramin resistance. By combination of a mapping-based approach with a de novo transcriptome assembly, a new variant surface glycoprotein, VSGsur, was identified. Subsequent experiments showed that the expression of VSGsur is enough to cause ~100-fold suramin resistance in T. brucei. The phenotypic changes in these parasites were not limited to drug resistance; the uptake of a number of different substrates and nutrients was highly reduced. This suggests on the one hand that the VSGsur-mediated resistance phenotype is caused by lower levels of intracellular suramin due to a reduced suramin uptake, linked to a decreased uptake of transferrin and low density lipoprotein. On the other hand, these results demonstrate that VSGs have an impact on the cell biology of trypanosomes that is broader than previously believed and reaching beyond immune evasion.
I further selected these trypanosomes for even higher levels of suramin resistance. Analysis of sequence variations revealed a non-synonymous mutation in the RuvB-like helicase. This mutation was absent in the sensitive parent clone, heterozygous after suramin selection for three months, and turned homozygous during the course of further selection. Even though this finding needs further experimental validation, we have strong indications that RuvB-like helicase is a suramin target in trypanosomes, since (i) an unrelated suramin-resistant T. evansi isolate bears a mutation of the same residue, and (ii) suramin was shown to inhibit helicases in viruses.
Taken together, the transcriptome analysis of suramin-resistant trypanosomes led to the identification of two new resistance mechanisms: A VSGsur-mediated resistance that causes a reduced suramin uptake; and a mutation in the RuvB-like helicase that might protect this potential drug target from suramin action.