Abstract
PURPOSE: The role of thiols in the reduction of Pt(IV) antitumor agents to Pt(II) is well recognized and it is widely thought that this reaction is required for activity. The sources of extracellular thiols in cell culture have been less studied. The purpose of the present work was to determine whether the stability of Pt(IV) complexes in culture medium can be affected by thiols that are released by cancer cells. METHODS: A two-column HPLC assay with UV/visible detection was used to determine the stability of two Pt(IV) complexes in culture medium with and without cells. The kinetics of the thiol release from a human ovarian cancer cell line SK-OV-3 and a human glioblastoma cell line U-87 MG were determined by a modification of the Ellman's method. RESULTS: The stability of a Pt(IV) complex with equatorial iodo ligands, trans, cis-[Pt(en)(OAc)(2)I(2)], was dramatically lower in culture medium in the presence of cells than in fresh culture medium, whereas the half-life of the dichloro analog, trans,cis-[Pt(en)(OAc)(2)Cl(2)], was somewhat increased. Although both complexes showed similar in vitro cell-growth-inhibitory activity, trans, cis-[Pt(en)(OAc)(2)Cl(2)] required a longer incubation time than the iodo analog to reach its maximal effect. The thiol content of the culture medium in the presence of cells was measured after 2 days: the concentrations from cultures of U-87 MG and SK-OV-3 cells were 3. 6 +/- 0.1 microM and 9.3 +/- 0.1 microM respectively, compared to 0. 07 +/- 0.04 microM in fresh medium. During the rapid growth phase, the extracellular thiol content reached a maximum of 20.0 +/- 0.5 microM and 47.8 +/- 0.2 microM for U-87 MG and SK-OV-3 cells respectively. CONCLUSIONS: These findings show that the culture medium conditioned by cancer cells can influence the stabilities of certain Pt(IV) complexes in cytotoxicity studies.