Synthesis and Characterization of (OC-6-33)-Dichloridobis((4-(13C2)ethoxy)-4-(13C4)oxobutanoato)bis((15N)ethylamine)platinum(IV). Time-Dependent NMR Spectroscopy and HPLC-MS Investigations on its Stability in Cell Culture Medium

Abstract

In order to investigate the stability of an experimental anticancer platinum(IV) prodrug in cell culture media, (OC-6-33)-dichloridobis((4-ethoxy)-4-oxobutanoato)bis(ethylamine)-platinum(IV) was synthesized with ¹⁵N labeled equatorial and fully ¹³C labeled axial ligands. The title compound as well as its stability in cell culture media was investigated in depth by ¹H, ¹³C, ¹⁵N, and ¹⁹⁵Pt one- and two-dimensional NMR spectroscopy as well as HPLC-MS. Different platinum(IV) compounds as a result of hydrolysis were detected in cell culture media as main species. Besides hydrolysis of one or both equatorial chlorido ligands, also hydrolysis of the ester moiety in the axial ligands and release of ethanol could be observed. The different mono and dihydroxidoplatinum(IV) species were proven by ¹⁹⁵Pt and ¹³C NMR spectroscopy as well as HPLC coupled to MS and HRMS. Reduction of the title compound to the corresponding platinum(II) analog or reaction of the complex with ingredients of the cell culture media were not observed. In cell culture medium supplemented with fetal calf serum (FCS), the formation of hydrolysis species was slowed down by a factor of two.