X-ray fluorescence imaging of single human cancer cells reveals that the N-heterocyclic ligands of iodinated analogues of ruthenium anticancer drugs remain coordinated after cellular uptake.

Analogues of KP1019 containing iodinated indazole ligands were prepared to investigate the biological fate of the Ru-N-heterocycle bond in this class of anticancer agents. The new complexes, 5-iodoindazolium trans-tetrachloridobis(5-iodoindazole)ruthen(III)ate (1) and 5-iodoindazolium trans-tetrachlorido(dimethyl sulfoxide)(5-iodoindazole)ruthen(III)ate (3), were characterized by elemental analysis, mass spectrometry and UV-vis spectrophotometry. Tetramethylammonium salts of these complexes (2 and 4) were synthesized and characterized in a similar manner. Half-maximum inhibitory concentrations of 2 and 4 with regard to A549 cells at 24 h were determined on the basis of the dose-response curves derived from real-time cell adhesion impedance measurements and were shown to be in the same range as those determined for KP1019 and NAMI-A using the same method. X-ray fluorescence imaging of single cultured A549 cells treated with 2 or 4 showed that, in both cases, the distribution of ruthenium and iodine was identical, indicating that the Ru-N bonds in the anionic complexes remained intact after incubation in culture medium and subsequent cellular uptake and processing.

Distinct cellular fates for KP1019 and NAMI-A determined by X-ray fluorescence imaging of single cells.

Small molecule ruthenium complexes show great promise as anticancer pharmaceuticals, but further rational development of these as drugs is stymied by an incomplete understanding of the mechanisms that give rise to markedly different biological behaviour for structurally similar species. X-ray fluorescence imaging at two incident energies was used to reveal the intracellular distribution of Ru in single human cells treated with KP1019, showing Ru localised in both cytosol and in the nuclear region. In addition the imaging showed that treatment with KP1019 modulated Fe distribution to resemble the Ru distribution, without affecting cellular Fe content. In stark contrast, Ru could not be visualised in cells treated with NAMI-A, indicating that it was not internalised and supporting the proposition that its activity is exerted through a membrane-binding mechanism.