Indazolium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] (1, KP1019) and its analogue sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] (2, KP1339) are promising redox-active anticancer drug candidates that were investigated with X-ray absorption near edge structure spectroscopy. The analysis was based on the concept of the coordination charge and ruthenium model compounds representing possible coordinations and oxidation states in vivo. 1 was investigated in citrate saline buffer (pH 3.5) and in carbonate buffer (pH 7.4) at 37 °C for different time intervals. Interaction studies on 1 with glutathione in saline buffer and apo-transferrin in carbonate buffer were undertaken, and the coordination of 1 and 2 in tumor tissues was studied too. The most likely coordinations and oxidation states of the compound under the above mentioned conditions were assigned. Microprobe X-ray fluorescence of tumor thin sections showed the strong penetration of ruthenium into the tumor tissue, with the highest concentrations near blood vessels and in the edge regions of the tissue samples.

X-ray absorption spectroscopy (XAS) and micro-synchrotron based X-ray fluorescence (micro-SXRF) are element specific spectroscopic techniques and have been proven to be valuable tools for the investigation of changes in the chemical environment of metal centres. XAS allows the determination of the oxidation state, the coordination motif of the probed element, the identity and the number of adjacent atoms and the absorber-ligand distances. It is further applicable to nearly all types of samples independent of their actual physical state (solid, liquid, gaseous) down to μM concentrations. Micro-SXRF can provide information on the distribution and concentration of multiple elements within a sample simultaneously, allowing for the chemical state of several elements within subcellular compartments to be probed. Modern third generation synchrotrons offer the possibility to investigate the majority of the biologically relevant elements. The biological mode of action of metal-based compounds often involves interactions with target and/or transport molecules. The presence of reducing agents may also give rise to changes in the coordination sphere and/or the oxidation state. XAS and micro-SXRF are ideal techniques for investigating these issues. This tutorial review introduces the use of XAS and micro-SXRF techniques into the field of inorganic medicinal chemistry. The results obtained for platinum, ruthenium, gallium, gold and cobalt compounds within the last few years are presented.

Tris(8-quinolinolato)gallium(III) (1, KP46) is a very promising investigational anticancer drug. Its interaction with serum proteins, elemental distribution, and coordination in tissue were investigated with X-ray absorption (XAS) methods. Model compounds with mixed O, N, and/or S donor atoms are reported. The coordination and structure of 1 in cell culture medium (minimum essential medium, MEM) and fetal calf serum (FCS) were probed by XANES and EXAFS. The interaction of 1 with the serum proteins apotransferrin (apoTf) and human serum albumin (HSA) was addressed as well. By application of micro-XAS to tissue samples from mice treated with 1, the gallium distribution pattern was analyzed and compared to those of physiological trace elements. The complex 1 turned out to be very stable under physiological conditions, in cell culture media and in tissue samples. The coordination environment of the metal center remains intact in the presence of apoTf and HSA. The gallium distribution pattern in tumor and liver tissue revealed high similarities to the distribution patterns of Zn and Fe, minor similarities to Cu and Ni, and no similarity to Ca.

Ruthenium complexes are promising candidates for anticancer agents, especially NKP-1339 (sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)]), which is on the edge to clinical applications. The anticancer mechanism seems to be tightly linked to the redox chemistry but despite progress in human clinical trials the in vivo Ru oxidation state and the coordination of Ru remains unclear. The Ru-based anticancer drug NKP-1339 was studied applying XANES (Cl K- and Ru L2,3-edges) in tumor, kidney and liver tissue of a SW480 bearing mouse. Based on coordination charge and 3D XANES plots containing a series of model compounds as well as pre-edge analysis of the ligand Cl K-edge it is suggested that NKP-1339 remains in its +III oxidation state after 24 hours and at least one of the four chlorido ligands remain covalently bound to the Ru ion showing a biotransformation from RuIIIN2Cl4 to RuIIIClx(N/O)6−x (X = 1 or 2).