Redox Proteins GSTs A1, M1, O1, O2, P1; Prx-3, and Trx-1 Association with In Vitro Imexon Sensitivity in Multiple Myeloma Cell Lines.

Abstract

Multiple myeloma (MM) is an incurable disease that is characterized by the proliferation of clonal plasma cells in the bone marrow. Conventional chemotherapy and autologous stem cell transplantation have improved survival. However, because MM remains an incurable disease, innovative approaches are needed. Imexon is a small-molecule, aziridine-containing iminopyrrolidone with previously demonstrated cytotoxic activity in MM. It acts bybinding to sulfhydryls, decreasing glutathione levels,increasing reactive oxygen species (ROS),changing mitochrondial membrane potential,releasing cytochrome C into the cytoplasm, andactivating caspases and thus inducing apoptosis.Phase I clinical trials have shown that imexon is relatively nontoxic and can induce responses in patients with diverse malignancies. Imexon has shown antitumor activity in vitro against a variety of tumors, with MM cell lines and MM fresh tumor cells having the highest sensitivity. Imexon is currently being studied in a phase I/II MM clinical trial. Since imexon depletes glutathione, we determined if the following basal levels of oxidation-reduction (redox) proteins would be predictive of sensitivity: glutathione S-transferases (GSTs) A1, M1, O1, O2, P1; peroxiredoxin-3 (Prx-3), and thioredoxin-1 (Trx-1). Imexon showed single agent activity against MM cell lines by XTT cell proliferation assays, consistent with previous reports (see Table). By Western blotting, the GSTs A1 and M1 were expressed at low levels, which did not seem to correlate with imexon sensitivity. GST P1 was expressed differentially in MM cell lines, but without an obvious correlation to imexon sensitivity. GST O1 and O2 showed modestly increased protein levels in the resistant 8226/IM10 cell line compared to the sensitive 8226/S cells. Prx-3 and Trx-1 were uniformly expressed except for the very low or absent expression of Trx-1 in MM-1 cells. While basal levels of most of the redox proteins tested were not predictive of imexon sensitivity, changes in redox protein levels after drug exposure may be predictive and will be tested in future studies. Additionally, other redox proteins may be better predictive markers. In conclusion, imexon is active as a single agent in vitro in MM cell lines. Basal levels of the redox proteins GST A1, M1, P1; Prx-3, and Trx-1 do not appear to correlate with imexon sensitivity in the cell lines tested. GST O1 and O2 are expressed at higher levels in the sensitive 8226/S cell line, suggesting potential utility as predictive markers. Future studies evaluating changes in redox protein levels after drug exposure in MM cell lines and in MM patients in an ongoing imexon clinical trial may further validate GST O1 and O2 as well as provide additional predictive molecular markers.Redox Protein Basal Levels in MM Cell Lines by Western BlotMM Cell Line (IC50, uM)GST A1GST M1GST O1GST O2GST P1Prx-3Trx-1OCI (74)++++++++++++++++8226/S (95)−+/−+++++++++MM-1 (180)++/−++++++++−8226/IM10 (270)++/−++++++/−+++++U266/1300 (528)++/−+++++++++++