Reaction kinetics and cell effects of a new glutathione peroxidase mimetic drug: PhSeZnCl

The glutathione (GSH)-dependent reduction reaction of H2O2 and lipid hydroperoxides catalyzed by the enzyme glutathione peroxidase (GPx), has a critical role in tissue protection and cell signaling. Our laboratories have recently synthesized a series of Selenium-organic drugs with anticipated GPx mimetic activity, of which PhSeZnCl is one of the lead compounds. In this study, the catalytic mechanism and potency of this GPx-mimetic drug were assessed in the GSH-reductase (GR)-coupled assay. Comparisons with the test drugs PhSe)2, an oxidized form of PhSeZnCl, and the prototypical organo-Se drug with GPx activity Ebselen, were carried out. Preliminary experiments were made varying the concentration of the catalysts, keeping steady the co-substrates H2O2 and GSH (0.21 and 1.0 mM, respectively). Under these conditions, the reaction kinetics for all the test compounds was of the same order up to a concentration of the catalysts of 20 μM. Kinetics data obtained varying the concentrations of the co-substrates and maintaining the catalysts at the final concentration of 10 µM revealed that PhSeZnCl is a more potent GPx mimetic drug than Ebselen and PhSe)2, showing slightly higher Vmax then Ebselen and excellent catalytic efficiency (kcat/KM) with lower KM values for both the two substrates H2O2 and GSH, in comparison with the other catalysts. Given the high thiol reactivity of PhSeZnCl, a microencapsulated formulation (M-PhSeZnCl) was prepared by spray dried technology using D,L-lactate as coating polymer. Microencapsulation resulted in a lower GPx-mimetic activity and catalytic efficiency, but significantly lowered cytotoxicity as assessed in several cell model systems. Uptake and redox effects of the microencapsulated formulation of PhSeZnCl were assessed in MCF-7 cells. M-PhSeZnCl slightly affected resting level of intracellular ROS and significantly lowered the burst of cellular ROS induced by a challenge with 1mM extracellular H2O2 in cells expressing allelic variants of GSTP1-1 with higher activity in the restoration of the membrane peroxidase and signaling protein peroxiredoxin 6. The findings in this study show the potential of PhSeZnCl as GPx mimetic drug and selective redox-active agent in cell treatments.