Anti-parasitic dinuclear thiolato-bridged arene ruthenium complexes alter the mitochondrial ultrastructure and membrane potential in Trypanosoma brucei bloodstream forms

Abstract

Trypanosoma brucei causes human African trypanosomiasis and Nagana disease in cattle, imposing substantial medical and economic burden in sub-Saharan Africa. The current treatments have limitations, including the requirement for elaborated protocols, development of drug resistance, and they are prone to adverse side effects. In vitro screening of a library of 14 dinuclear-thiolato bridged arene ruthenium complexes, originally developed for treatment of cancer cells, resulted in the identification of 7 compounds with IC50 values ranging from 3 to 26 nM. Complex [(η6-p-MeC6H4Pri)2Ru2(μ2-SC6H4-o-Pri)3]Cl (2) (IC50 = 4 nM) and complex [(η6-p-MeC6H4Pri)2Ru2(μ2-SCH2C6H4-p-But)2(μ2-SC6H4-p-OH)]BF4(9) (IC50 = 26 nM) were chosen for further assessments. Application of complex 2 and 9 at 20 nM and 200 nM, respectively, for 4.5 h induced alterations in the trypanosome mitochondrion as evidenced by immunofluorescence employing an antibody against mitochondrial Hsp70 and Mitotracker labeling. Transmission electron microscopy of parasites taken at 2 and 4h of treatment demonstrated massive alterations in the mitochondrial ultrastructure, while other organelles and structural elements of the parasites remained unaffected. Complex 2 treated trypanosomes exhibited a distorted mitochondrial membrane, and the mitochondrial matrix was transformed into an amorphous mass with different degrees of electron densities. Complex 9 did not notably impair the integrity of the membrane, but the interior of the mitochondrion appeared either completely translucent, or was filled with filamentous structures of unknown nature. Dose- and time-dependent effects of these two compounds on the mitochondrial membrane potential were detected by tetramethylrhodamine ethyl ester assay. Thus, the mitochondrion and associated metabolic processes are an important target of dinuclear thiolato-bridged arene ruthenium complexes in T. brucei.