Activation of EGFR by small compounds through coupling the generation of hydrogen peroxide to stable dimerization of Cu/Zn SOD1

Abstract

Activation of cell signaling by reactive chemicals and pollutants is an important issue for human health. It has been shown that lipophilic nitro-benzoxadiazole (NBD) compounds rapidly move across the plasma membrane and enhance Epidermal Growth Factor Receptor (EGFR) tyrosine phosphorylation in cancer cells. Unlike ligand-dependent activation, the mechanism of this induction relies on the generation of hydrogen peroxide, which is involved in the activation of the catalytic site of the receptor and the inactivation of protein tyrosine phosphatase PTP-1B. Production of H2O2 during redox transformation of NBD compounds is associated with the transition of a monomeric form of Cu/Zn superoxide dismutase 1 (SOD1) to stable dimers. The highly stable and functionally active SOD1 dimer, in the absence of adequate activities in downstream reactions, promotes the disproportionate production and accumulation of intracellular hydrogen peroxide shortly after exposure to NBD compounds. The intrinsic fluorescence of small compounds was used to demonstrate their binding to SOD1. Our data indicate that H2O2 and concomitantly generated electrophilic intermediates behave as independent entities, but all contribute to the biological reactivity of NBD compounds. This study opens a promising path to identify new biomarkers of oxidative/electrophilic stress in the progression of cancer and other diseases.

Reactive Chemicals and Electrophilic Stress in Cancer

“…On the contrary, a significant proportion of Cu/Zn SOD1, which is mainly located in the cytoplasm and much less in mitochondria [24], was detected as a 32-kDa dimer in cells exposed to lipophilic NBD compounds for 5-10 min. The induced dimers of SOD1 were stable and they did not disrupt during electrophoretic migration in a denaturing gel in contrast to less stable dimers of native enzyme in non-treated cells [22].…”

“…Generation of intracellular ROS by NBD compounds may be related to the action of superoxide dismutase, catalase, and/or glutathione peroxidase, which are responsible for the protection of human cells against toxic oxygen species [21]. Given that the activity of these enzymes is exhibited by dimeric or higher oligomeric forms, we studied the oligomerization state of corresponding proteins in breast cancer and prostate cancer cells exposed to NBD compounds [22,23]. No changes were detected in the oligomerization or expression of catalase and glutathione peroxidase.…”

Discovery of compounds inhibiting the ADP-ribosyltransferase activity of pertussis toxin

“…In part, this could relate to the fact that NSC228155 easily permeates cells. It has been shown in MDA-MB-468 breast cancer cells that 5 min incubation with 100 µM NSC228155 caused rapid movement of this inherently fluorescent molecule across cell membranes and dispersal to both the cytoplasm and nucleus 29 . However, we witnessed significant toxicity of NSC228155 for HEK293T cells with 20 µM or higher concentrations after a 3.5-h incubation (cytotoxicity IC 50 – 15.33 µM see Fig.…”

“…6E). This cytotoxicity might be caused by the proposed NSC228155-mediated production of reactive oxygen species (ROS) inside the cell 29 . In this respect, it is noteworthy that, upon incubation of cells with 5 µM of NSC228155 for 2.5 h, we did not detect auto-ADP-ribosylation of PARP1/ARTD1 (see Fig.…”

α,1,6-Fucosyltransferase (FUT8) regulates cancer-promoting capacities of cancer-associated fibroblasts (CAFs) through the modification of EGFR core fucosylation in non-small cell lung cancer

“…Lens culinaris agglutinin, a lectin that can specifically identify fucose, was purchased from Vector Labs (California, USA). NSC 228155, an activator of EGFR which binds to the extracellular region of EGFR and enhance tyrosine phosphorylation of EGFR [48] , was purchased from MedChemExpress (New Jersey, USA). Other chemicals and reagents were purchased from Sigma (St. Louis, USA) and Vetec (St. Louis, USA).…”