Nitroxides Tempol and Tempo Induce Divergent Signal Transduction Pathways in MDA-MB 231 Breast Cancer Cells

Abstract

Tempol and tempo are stable free radical nitroxides that possess antioxidant properties. In this study, we examined the effects of these compounds on components of the mitogen-activated protein kinase signal transduction cascade. Tempo treatment (15 min) of MDA-MB 231 human breast cancer cells resulted in significant levels of tyrosine phosphorylation of several as yet unidentified proteins compared with equimolar concentration of tempol (10 mM). Both compounds caused tyrosine phosphorylation and activation of Raf-1 protein kinase (30 min, 2-3-fold). Interestingly, however, only tempol caused increased extracellular signal-regulated kinase 1 activity (2 h, ϳ3-fold). On the other hand, tempo, but not tempol, potently activated stress-activated protein kinase (2 h, >3-fold). Consistent with these data, tempol was found to be noncytotoxic, whereas tempo induced apoptotic cell death (2 h, >50%). Tempo treatment also resulted in significant elevation of ceramide levels at 30 min (54% over control) and 1 h (71% over control) posttreatment, preceding stress-activated protein kinase activation and apoptosis. These data suggest that in the absence of an environmental oxidative stress, tempol and tempo elicit distinct cellular signaling pathways. The recognition of the molecular mechanisms of nitroxide action may have important implications for biological effectiveness of these compounds.

Nitroxide TEMPO: A genotoxic and oxidative stress inducer in cultured cells

“…20% (Table 1), and 2.3 mM resulting in essentially no growth of TK6 cells (Table 2). These result are consistent with a time course analysis of TEMPO cytotoxicity in different types of tumor cells, which indicated that TEMPO treatment resulted in a > 50% decrease in the number of viable cells and a steady increase in apoptosis during the first 2 h of exposure, followed by a considerable increase in necrosis by 3 h (Suy et al, 1998). A 24 h treatment with TEMPO was cytotoxic to HaCaT human keratinocytes, producing an IC 50 of 2.66 mM using the amido-black-assay (Kroll et al, 1999).…”

“…A 24 h treatment with TEMPO was cytotoxic to HaCaT human keratinocytes, producing an IC 50 of 2.66 mM using the amido-black-assay (Kroll et al, 1999). It has been reported that TEMPO is approximately 200 times more lipophilic than its derivative 4-hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl (4-hydroxy-TEMPO) (Suy et al, 1998), which may facilitate its accumulation in the cell membrane and potentiate its cellular toxicity. In addition, TEMPO is a unique thermosensitizer that synergistically induces cell death when 5-10 mM TEMPO is presented during heating at 44°C in U937 leukemic cells and results in Bax activation and mitochondrial outer membrane permeabilization (Zhao et al, 2010).…”

A Mitochondria-Targeted Nitroxide/Hemigramicidin S Conjugate Protects Mouse Embryonic Cells Against Gamma Irradiation

“…Additionally, 5-125 may act as an SOD-mimetic and dismutate superoxide radicals to hydrogen peroxide (24,25). Finally, Suy et al (44) reported that the nontargeted nitroxide, TEMPOL, stimulated the extra cellular signal-regulated kinase (ERK) signaling pathway, which primarily promotes growth and proliferation/survival.…”

High-Resolution Mapping of Tumor Redox Status by Magnetic Resonance Imaging Using Nitroxides as Redox-Sensitive Contrast Agents

“…Nitroxides provide protection in cells and tissues against diverse types of insult, such as exposure to superoxide and hydrogen peroxide, by directly scavenging the ROS (31). In addition to protection by radical scavenging, nitroxides also evoke cellular responses such as cell signaling -specific pathways (32,33). The ability to trigger cell signaling -specific pathways has been postulated to support possible antitumor effects of nitroxides in vivo (34,35).…”