NBMI Exerts Protection Against Copper Overload in HepG2 and SH‐SY5Y Cell Lines

Abstract

Copper (Cu) is a naturally occurring essential element required for many biological processes. At low concentrations, Cu plays a vital role by getting incorporated into various metalloenzymes and at higher concentrations can be harmful to hepatic, central, and renal systems. Chronic toxicity with high concentrations of Cu have been characterized in Wilsons’s disease patients, and chelation has been the mainstay of therapy. As the currently approved chelator D‐pencillamine is associated with many adverse effects, the search for other therapeutic interventions is ongoing. N,N′bis‐(2‐mercaptoethyl) isophthalamide (NBMI) is a novel metal chelator and thiol redox antioxidant. NBMI has been shown to chelate harmful heavy metals such as mercury and lead in vitro. We explored whether NBMI displayed cytoprotective effects in HepG2 and SH‐SY5Y cells exposed to copper sulfate (CuSO4). Both the cell types were treated with either vehicle or NBMI (0 ‐ 100 μM) for 24 hours followed by CuSO4 (750 μM) for an additional 24 hours. Cell viability (CellTiter‐Glo® Assay) and degree of oxidative stress (ROS Assay) were determined. Comparisons between multiple groups were performed using analysis of variance (ANOVA) followed by a post hoc Tukey’s test. Results with p‐value <0.05 were considered statistically significant. Exposure of HepG2 and SH‐SY5Y cells to CuSO4 decreased cell viability in a concentration‐dependent manner. In addition, CuSO4 exposure led to an increase in ROS levels in HepG2 cells indicating oxidative stress. Pretreatment with NBMI diminished CuSO4‐induced cell death in both cell types and decreased ROS levels in HepG2 cells. The results imply that NBMI is a new and promising chelator for treating Cu‐overload genetic disorders.