In general, ruthenium complexes 1 and 3 show a higher inhibitory potency on Cdk2/cyclin E than their osmium congeners
2 and 4. At a concentration of 10 μM, ruthenium complexes 1 and 3 yield 43% and 37% inhibition, which is about twice as high as the effect exerted by osmium congeners 2 and 4. A 50% inhibition of Cdk2/cyclin E requires concentrations buy Ruxolitinib of up to 40 μM (or even higher in the case of 2) (Fig. 3). Correlation with cytotoxic potencies is rather weak overall, but closest at the intermediate concentration of 10 μM. Given the capacity of inhibiting Cdk activity, an impact on the cell cycle of proliferating cells might be expected from these compounds. Therefore, changes in cell cycle distribution induced by 1–4 were studied in exponentially growing A549 cells treated with these learn more compounds in varying concentrations for 24 h, then stained with propidium iodide and analyzed for their DNA content by flow cytometry.
The compounds 1–4 have only weak effects on the cell cycle within the concentration range tested (Fig. 4). A slight increase of the G0/G1 fraction and a decrease of the S phase fraction could be observed up to a concentration of 40 μM of complexes 1 and 2. Reduced numbers of cells in G2/M phase compared to the control are visible at low concentrations of these compounds (2.5 μM and 10 μM). In the case of complexes 3 and 4, the cell fraction in G0/G1 phase is slightly increased only at the lowest (2.5 μM) and/or the medium concentration
(10 μM) of the compounds. The inhibitory potency of the ruthenium and osmium complexes on DNA synthesis was determined by the BrdU assay. All four compounds inhibit BrdU incorporation into DNA of A549 non-small cell lung cancer cells within 24 h. Although the compounds have little effect on the cell cycle, a clear reduction of DNA synthesis could be observed (Fig. 5). Ruthenium complexes 1 and 3 are again Cediranib (AZD2171) somewhat more effective than the corresponding osmium complexes 2 and 4, in accordance with the structure–activity relationships revealed in the MTT assay. A concentration of 5 μM resulted in nearly 50% and 30% inhibition of BrdU incorporation by 1 and 3, respectively, whereas the effects of 2 and 4 are still modest. In any case, a strong reduction of DNA synthesis requires concentrations higher than 5 μM. A concentration of 20 μM, however, is sufficient for diminishing BrdU incorporation to values below 15% for all compounds. Cellular accumulation of complexes 1 and 3 was studied in the colon carcinoma cell line SW480. The cells were incubated at 37 °C for 2 h with 10 μM of the respective compound, and cellular metal contents were then determined by ICP-MS measurement, revealing that cellular amounts of ruthenium are one third lower after exposure to 1 (2.0 ± 0.3 fmol/cell) than those after treatment with 3 (3.0 ± 0.2 fmol/cell). These results do not correlate with cytotoxicity (compare Fig. 2b).