In Vitro Study of the Enzymatic and Nonenzymatic Conjugation of Treosulfan with Glutathione

Abstract

Background and ObjectivesTreosulfan (dihydroxybusulfan), licensed for the treatment of ovarian carcinoma, is investigated in clinical trials as a myeloablative agent for conditioning prior to hematopoietic stem cell transplantation. Clinical experience shows that treosulfan exhibits lower organ toxicity than busulfan, including hepatotoxicity. Elimination of busulfan primarily via enzymatic conjugation with glutathione (GSH) in the liver is considered to be the main cause of the drug’s hepatotoxicity and interpatient clearance variability. It is believed that treosulfan undergoes no hepatic metabolism but empirical evidence is lacking. The aim of this kinetic study was to verify if treosulfan is capable of conjugating with GSH.MethodsTreosulfan (200 μM) was incubated at pH 7.2 and 37 °C with 5 mM GSH in the presence or absence of human liver cytosol, the main store of glutathione S-transferase in the body. Concentrations of treosulfan were determined using liquid chromatography–tandem mass spectrometry and then subjected to kinetic analysis.ResultsThe decay of treosulfan in the solution followed a one-exponential model in the presence of either GSH or liver cytosol and GSH. The first-order reaction rate constants (0.25 h−1) did not differ statistically from those found for treosulfan conversion in pH 7.2 buffer only.ConclusionTreosulfan does not undergo either spontaneous or enzymatic conjugation with GSH at a noticeable rate. The result indicates that the clearance of treosulfan is independent of glutathione S-transferase activity, GSH stores, and co-administration of drugs utilizing the GSH metabolic pathway.

Novel insights into conjugation of antitumor-active unsymmetrical bisacridine C-2028 with glutathione: Characteristics of non-enzymatic and glutathione S-transferase-mediated reactions

“…These can occur spontaneously, proceed with the participation of small-molecule compounds or be light-induced. As has been reported for several compounds (i.e., isothiocyanates or catecholamines), the reactive GSH thiol-group, which possesses strong nucleophilic properties, provides the redox potential also for non-enzymatic conjugation [ 10 , 29 , 30 ].…”

“…A significant route of drug detoxification in many species is the conjugation of xenobiotic with reduced tripeptide glutathione (GSH; L- γ -glutamyl-L-cysteinyl-glycine) [ 8 ]. Due to the high nucleophilic reactivity of the GSH thiol group, this process can be non-enzymatic and/or mediated by various GSH-dependent enzymes [ 9 , 10 ]. On the other hand, depending on the properties of the substrate, GSH conjugation may also play an important role in bioactivation reactions as it can generate toxic conjugates [ 11 ].…”

Treosulfan Exposure Predicts Thalassemia‐Free Survival in Patients with Beta Thalassemia Major Undergoing Allogeneic Hematopoietic Cell Transplantation

“…Patients with GSTA1*B are poor metabolizers of busulfan, thereby exhibiting high systemic busulfan exposure, leading to organ toxicities and adverse HCT outcomes 40–43 . Romański et al 44 . performed a kinetic analysis to examine treosulfan–GSH conjugation in vitro and showed that treosulfan does not undergo spontaneous or GST‐mediated conjugation with GSH.…”

“…Patients with GSTA1*B are poor metabolizers of busulfan, thereby exhibiting high systemic busulfan exposure, leading to organ toxicities and adverse HCT outcomes. [40][41][42][43] Romański et al 44 performed a kinetic analysis to examine treosulfan-GSH conjugation in vitro and showed that treosulfan does not undergo spontaneous or GST-mediated conjugation with GSH. We screened for GSTA1*B haplotype, which was shown to explain variability in busulfan PK in the present study, and assessed the effect of the genotype on treosulfan/S, S-EBDM exposure.…”