Metal-based anticancer agent development can be improved with advanced metallomics methods that allow for quick and efficient screening of metallodrugs for their metabolites in biological media. Cellular accumulation in in vitro settings is not always correlated with cytotoxicity; and protein binding, particularly with albumin and transferrin, can have an important influence on metallodrug transportation, selectivity, and efficacy. We contrast the time-dependent cellular accumulation of both cisplatin and the pre-clinically investigated RAPTA-C in terms of cell uptake and speciation in culture medium via CE-ICP-MS analysis. Despite RAPTA-C being administered at 40-fold higher dose than cisplatin, owing to its much higher IC value, the accumulation over time was only 10-fold higher. An optimised CE-ICP-MS method, through the coating of the capillary to prevent protein-capillary surface interactions, resulted in superior resolution and metal-protein adduct identification. It was then used for extracellular speciation in conjunction with [tris(acetylacetonato)cobalt(iii)] as an internal standard. RAPTA-C was found to be more inert to extracellular reactions than cisplatin which could be used to rationalise the observed cellular uptake patterns. While for cisplatin both transferrin and albumin were identified as the main binding partners, RAPTA-C was found to react nearly exclusively with albumin. Moreover, this behaviour was time-dependent and our results also demonstrate that cancer cells have an influence on metal species distribution in the cell culture medium over time.
Run-by-run variations are very common in capillary electrophoretic (CE) separations and cause imprecision in both the migration times and the peak areas. This makes peak and kinetic trend identification difficult and error prone. With the aim to identify suitable standards for CE separations which are compatible with the common detectors UV, ESI-MS, and ICP-MS, the Co complexes [Co(en)]Cl, [Co(acac)] and K[Co(EDTA)] were evaluated as internal standards in the reaction of the anticancer drug cisplatin and guanosine 5'-monophosphate as an example of a classical biological inorganic chemistry experiment. These Co chelate complexes were considered for their stability, accessibility, and the low detection limit for Co in ICP-MS. Furthermore, the Co complexes are positively and negatively charged as well as neutral, allowing the detection in different areas of the electropherograms. The background electrolytes were chosen to cover a wide pH range. The compatibility to the separation conditions was dependent on the ligands attached to the Co centers, with only the acetylacetonato (acac) complex being applicable in the pH range 2.8-9.0. Furthermore, because of being charge neutral, this compound could be used as an electroosmotic flow (EOF) marker. In general, employing Co complexes resulted in improved data sets, particularly with regard to the migration times and peak areas, which resulted, for example, in higher linear ranges for the quantification of cisplatin.
Cisplatin and its second and third generation analogues are widely used in the treatment of cancer. To study their reactions with proteins, we present a method based on SDS‐PAGE separation and laser ablation–inductively coupled plasma‐mass spectrometry (LA–ICP‐MS) for platinum detection in the reaction between human serum albumin (HSA) and cisplatin. We developed matrix‐matched standards of HSA/cisplatin mixtures and used them to quantify the amount of adducts formed at different HSA:cisplatin ratios. We noted that cisplatin incubation with HSA resulted in the formation of higher order HSA n‐mers, depending on the amount of cisplatin added. This caused a depletion of the HSA dimer bands, while the majority of HSA was present as the monomer. Inducing the formation of such higher molecular weight species may have an impact on the mode of action of metallodrugs.
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