Dithiolethiones D3T and ACDT Protect Against Iron Overload-Induced Cytotoxicity and Serve as Ferroptosis Inhibitors in U-87 MG Cells

Abstract

Iron overload-induced oxidative stress is implicated in various neurodegenerative disorders. Given the numerous adverse effects associated with current iron chelators, natural antioxidants are being explored as alternative therapeutic options. Dithiolethiones found in cruciferous vegetables have emerged as promising candidates against a wide range of toxicants owing to their lipophilic and cytoprotective properties. Here, we test the dithiolethiones D3T and ACDT against ferric ammonium citrate (FAC)induced toxicity in U-87 MG astrocytoma cells. Exposure to 15 mM FAC for 24 hours resulted in 54% cell death. A 24-hour pretreatment with 50 µM D3T and ACDT prevented this cytotoxicity. Both dithiolethiones exhibited antioxidant effects by activating the Nrf2 transcription factor and upregulating levels of intracellular glutathione (GSH). This resulted in the successful inhibition of FAC-induced reactive oxygen species, lipid peroxidation, and cell death. Additionally, D3T and ACDT upregulated expression of the Nrf2-mediated iron storage protein ferritin which consequently reduced the total labile iron pool. A 24-hour pretreatment with D3T and ACDT also prevented cell death induced by the ferroptosis inducer erastin by upregulating the xCT transporter expression. The resulting increase in intracellular GSH and alleviation of lipid peroxidation was comparable to that caused by ferrostatin-1, a speci c ferroptosis inhibitor. Collectively, our ndings demonstrate that dithiolethiones may show promise as potential therapeutic options for the treatment of iron overload disorders.

Ferroptosis inhibitors: past, present and future

“…Upregulates HO-1, GPX4 expression and cellular activity Upregulation of GPX4, SLC7A11 levels and downregulation of p53 expression (Chen et al, 2021c;Zhai et al, 2022;Zhai et al, 2023b) " neutrophil piggybacking" strategy Neutrophil therapy in combination with Se targets delivery of antioxidant enzymes and upregulates GPX4 (Wang et al, 2022a;Xu et al, 2024) Frontiers in Pharmacology frontiersin.org while also protecting U-87MG cells from iron overload-induced cytotoxicity (Kulkarni et al, 2023). Additionally, D3T upregulates silent information regulator 1 (SIRT1), Nrf2, and HO-1, ameliorating cognitive deficits in the Tg2576 AD mouse model (Cui et al, 2018).…”

“…Dithiolethiones, lipophilic organosulfur compounds ( Ansari et al, 2018 ), such as 3H-1,2-dithiole-3-thione (D3T) and 5-amino-3-thioxo-3H-(1,2)dithiole -4-carboxylic acid ethyl ester (ACDT) , activate Nrf2 and increase system Xc- and GSH levels in the erastin-induced ferroptosis model, demonstrating antioxidant effects and significant inhibition of ferroptosis akin to Fer-1 ( Kulkarni et al, 2023 ). Treatment with D3T and ACDT upregulated Nrf2-mediated expression of ferritin and FPN1, while also protecting U-87MG cells from iron overload-induced cytotoxicity ( Kulkarni et al, 2023 ). Additionally, D3T upregulates silent information regulator 1 (SIRT1), Nrf2, and HO-1, ameliorating cognitive deficits in the Tg2576 AD mouse model ( Cui et al, 2018 ).…”