Glutathione conjugation of the antitumor-active 1-nitroacridine derivatives compounds C-857 and C-1748 – the major role of glutathione S-transferase M1-1

Objectives: C-857 and C-1748 are antitumor-active agents, monomers of unsymmetrical bisacridine derivatives. The aim of this study was to analyze their glutathione (GSH) conjugation in vitro in the presence of glutathione S-transferase (GST) M1-1.
Methods: 0.01 mg/mL recombinant hGSTM1-1 (expressed in Escherichia coli, obtained from Sigma-Aldrich, MO, USA), 0.01 mM C-857 (C-1748), and 5 mM reduced L-glutathione (GSH) in 0.1 M potassium phosphate buffer solution (pH 6.5) were incubated at 37 °C for 0 and 60 min in a total volume of 50 µL. MgCl2 solution (2 mM) was added for enzyme stimulation. The incubations were terminated by adding ice-cold MeOH to the incubation mixtures (1:1, v/v). The samples were vortexed, placed in ice for 10 min, and centrifuged at 10000 x g for 10 min. Aliquots of the supernatants (50 µL) in triplicate were then analyzed directly by reversed-phase liquid chromatography (RP-LC) with UV-vis detection and/or diode array and multiple wavelength detection, and monitored by MS.
Key findings: Our data demonstrated that both C-857 and C-1748 are extensively metabolized by hGSTM1-1. It was possible to identify one metabolite whose initial structure was identified by LC/MS as a GSH monoconjugate of C-857 (C-1748). GST-mediated GSH conjugation of these compounds may play an important role in their elimination in humans, strongly affecting bioavailability.