Pentathiepins: A Novel Class of Glutathione Peroxidase 1 Inhibitors that Induce Oxidative Stress, Loss of Mitochondrial Membrane Potential and Apoptosis in Human Cancer Cells

Abstract

A novel class of glutathione peroxidase 1 (GPx1) inhibitors, namely tri‐ and tetracyclic pentathiepins, has been identified that is approximately 15 times more potent than the most active known GPx1 inhibitor, mercaptosuccinic acid. Enzyme kinetic studies with bovine erythrocyte GPx1 indicate that pentathiepins reversibly inhibit oxidation of the substrate glutathione (GSH). Moreover, no inhibition of superoxide dismutase, catalase, thioredoxin reductase or glutathione reductase was observed at concentrations that effectively inhibit GPx1. As well as potent enzyme inhibitory activity, the pentathiepins show strong anticancer activity in various human cancer cell lines, with IC 50 values in a low‐micromolar range. A representative tetracyclic pentathiepin causes the formation of reactive oxygen species in these cells, the fragmentation of nuclear DNA and induces apoptosis via the intrinsic pathway. Moreover, this pentathiepin leads to a rapid and strong loss of mitochondrial membrane potential in treated cancer cells. On the other hand, evidence for the induction of ferroptosis as a form of cell death was negative. These new findings show that pentathiepins possess interesting biological activities beyond those originally ascribed to these compounds.

The Effect of Glutathione Peroxidase-1 Knockout on Anticancer Drug Sensitivities and Reactive Oxygen Species in Haploid HAP-1 Cells

“…For the determination of the anticancer potency of various anticancer drugs on HAP-1 cells, the IC 50 values were determined with MTT cell viability assay as described previously [ 30 ]. Briefly, cells were seeded out in 96-well plates at a density of 5000 cells in 100 µL culture medium and allowed to attach for 24 h. Compounds were dissolved in either N,N -dimethylformamide (DMF; for the platinum compound) or dimethyl sulfoxide (DMSO; for all other compounds), then serial diluted at 1000-fold the intended concentration before being diluted 1:500 into culture medium.…”

“…For the determination of the ROS-content of cells after incubation with anticancer drugs, a cell cytometer based technique with the intracellular ROS sensor reagent DCF-DA was used as previously described [ 30 ]. Briefly, cells were seeded out in a density of 50,000 cells per 3 mL, into a 6-well plate and allowed to attach for 24 h. Then, cells were exposed to anticancer drugs in fractions of the IC 50 value and incubated for 24 h. After removal of the medium, cells were harvested with trypsin/EDTA and washed with PBS by a centrifugation and resuspension step, then treated with 500 µL of a 20 µM DCF-DA solution in PBS for 30 min in the dark.…”

Targeting the Redox Landscape in Cancer Therapy

“…A number of published studies show the involvement of GPx proteins in tumorigenesis and chemotherapy resistance, which is summarized in these reviews [195,196]. Currently, there is no selective inhibitor for GPx proteins for therapeutic application; however, recent developments in medicinal chemistry show promising advancements for targeting GPx1 [197] and GPx4 [198].…”

Thioredoxin and Glutaredoxin Systems as Potential Targets for the Development of New Treatments in Friedreich’s Ataxia

“…In addition, inhibition of glutathione peroxidase enzyme 4 (GPX4) [ 92 , 93 ] or GSH unavailability or defects in its restoration [ 88 , 94 ] produce lipid hydroperoxide accumulation that triggers ferroptosis. Importantly, the implication of TRX1 and TRXRD in ferroptosis has also been described [ 95 , 96 ]. Increased ROS, lower reduced GSH concentrations and enhanced sensitivity to oxidants compared with control neurons have also been observed in these FRDA cell models [ 97 , 98 ].…”