PMX464, a thiol-reactive quinol and putative thioredoxin inhibitor, inhibits NF-κB-dependent proinflammatory activation of alveolar epithelial cells

Background and purpose: Subtle changes in the intracellular reduction-oxidation (redox) state can modulate nuclear factor-kappa B (NF-kappa B) activity. Thioredoxin-1 (Trx) is a small, ubiquitous, redox-active thiol (-SH) protein that, with thioredoxin reductase-1 (TrxR), modifies the redox status of NF-kappa B pathway components. PMX464 is a novel thiol-reactive quinol thought to inhibit the Trx/TrxR system. The aim of this work was to investigate whether PMX464 inhibited NF-kappa B-mediated proinflammatory activation of human type II alveolar epithelial cells (A549). Experimental approach: Intercellular adhesion molecule-1 (ICAM-1), granulocyte-macrophage colony-stimulating factor (GM-CSF) and CXCL8, NF-kappa B DNA binding, nuclear translocation of NF-kappa B p65 subunit, I kappa Ba degradation, I kappa B phosphorylation and I kappa B kinase (IKK) activity were assessed in A549 cells stimulated with IL-1 beta with or without PMX464 pretreatment. Effects of PMX464 on ICAM-1 expression in human lung microvascular endothelial cells (HLMVEC) were also investigated. For comparison, selected measurements (ICAM-1 and I kappa B-alpha phospho-I kappa B-alpha) were made on A549 cells after RNA interference-mediated silencing (siRNA) of Trx. Key results: PMX464 reduced ICAM-1, GM-CSF and CXCL8 expression in IL-1 beta-stimulated A549 cells and ICAM-1 in HLMVEC. PMX464 inhibited IL-1 beta-induced NF-kappa B DNA binding, nuclear translocation of NF-kappa B p65 subunit and factors involved in NF-kappa B activation; specifically, IkBa degradation, I kappa B phosphorylation and I kappa B kinase (IKK) activity in A549. By contrast, Trx siRNA did not alter ICAM-1 expression or I kappa B alpha degradation/phosphorylation in IL-1 beta-stimulated A549 cells. Conclusion and implications: PMX464 inhibits a proinflammatory response in A549 cells targeting the NF kappa B pathway above IKK. The lack of effect with Trx siRNA suggests that PMX464 acts on thiol proteins, in addition to Trx, to elicit anti-inflammatory responses in lung epithelial cells.