ROS inhibitor N-acetyl-L-cysteine antagonizes the activity of proteasome inhibitors

Abstract

NAC (N-acetyl-l-cysteine) is commonly used to identify and test ROS (reactive oxygen species) inducers, and to inhibit ROS. In the present study, we identified inhibition of proteasome inhibitors as a novel activity of NAC. Both NAC and catalase, another known scavenger of ROS, similarly inhibited ROS levels and apoptosis associated with H2O2. However, only NAC, and not catalase or another ROS scavenger Trolox, was able to prevent effects linked to proteasome inhibition, such as protein stabilization, apoptosis and accumulation of ubiquitin conjugates. These observations suggest that NAC has a dual activity as an inhibitor of ROS and proteasome inhibitors. Recently, NAC was used as a ROS inhibitor to functionally characterize a novel anticancer compound, piperlongumine, leading to its description as a ROS inducer. In contrast, our own experiments showed that this compound depicts features of proteasome inhibitors including suppression of FOXM1 (Forkhead box protein M1), stabilization of cellular proteins, induction of ROS-independent apoptosis and enhanced accumulation of ubiquitin conjugates. In addition, NAC, but not catalase or Trolox, interfered with the activity of piperlongumine, further supporting that piperlongumine is a proteasome inhibitor. Most importantly, we showed that NAC, but not other ROS scavengers, directly binds to proteasome inhibitors. To our knowledge, NAC is the first known compound that directly interacts with and antagonizes the activity of proteasome inhibitors. Taken together, the findings of the present study suggest that, as a result of the dual nature of NAC, data interpretation might not be straightforward when NAC is utilized as an antioxidant to demonstrate ROS involvement in drug-induced apoptosis.

Piperlongumine (piplartine) and analogues: Antiproliferative microtubule-destabilising agents

“…No significant increases in cell viability were noted when cells were pre-treated with Trolox. Similar results show that pre-treatment with NAC but not Trolox has an effect on activity of 1 [19]. Compound 1 is a relatively reactive compound, due to the reactive α,β-unsaturated carbonyl system contained in the piperidinone ring.…”

“…Trolox (100 μM and 300 μM) has been shown to be an effective antioxidant in MDA-MB-231 breast cancer cells and MIA PaCa-2 pancreatic cancer cells [19]. We also evaluated a number of known tubulindestabilising agents [colchicine, 2 and four chalcones (14-17, Figure 5)].…”

“…Compound 1 has been recently reported to be a direct STAT3 inhibitor [4], a proteasome inhibitor [5], to promote autophagy [6], and to modulate NF-κB and NF-κB-regulated gene products [7].…”

Iron overloaded polarizes macrophage to proinflammation phenotype through ROS/acetyl‐p53 pathway

“…The results of Prussian staining are shown in Figure 1A, and showed that macrophages could rapidly take up iron ion. Considering that previous studies have shown a relationship between intracellular iron concentration and macrophage polarization,3, 5 we measured the expression of IL‐1β, IL‐10, TNF‐α, and TGF‐β with Western blotting (Figure 1B), the result showed expression of IL‐1β and TNF‐α increased. The results of qRT‐PCR and flow cytometry are shown in Figure 1C,D, and showed that both Fe 2+ and Fe 3+ induced the expression of M1‐associated maker such as TNF‐α, CD86, and iNOS.…”

“…ROS mainly include superoxide, hydrogen peroxide, singlet oxygen, ozone, hypohalous acids, and organic peroxides, all of which are byproducts of aerobic metabolism. Levels of ROS are mainly regulated by nonenzymatic antioxidants (eg, α‐tocopherol and glutathione) and antioxidant enzymes (eg, superoxide dismutase and catalase) 5. ROS physiologically regulates the synthesize of some kinds of proteins, such as phosphatase and tensin homologue, epidermal growth factor receptor, protein kinase B, and histone deacetylases,6, 7, 8, 9 and oxidizes DNA to promote transcription 4…”