Auranofin

Menadione (2-methyl-1,4-naphthoquinone sodium bisulfite, cat. No. M5750, Men) and sodium L-ascorbate (cat. No. A7631, Asc) were obtained from Sigma-Aldrich (St Louis, MO, USA); the reagents were dissolved in sterile distilled water. Auranofin (AUR) was purchased from Santa Cruz Biotechnology and dissolved in DMSO.

NADPH, 5,5-dithiobis (2-nitrobenzotic acid) (DTNB), glutathione reductase (GR, from Escherichia coli), dimethylsulfoxide (DMSO), 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT), bovine serum albumin, and digitonin were obtained from Sigma-Aldrich Corporation (St. Louis, MO, USA). Auranofin was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). Trypsin, penicillin, streptomycin, RPMI 1640 medium, and fetal calf serum were products of HyClone (Logan City, UT, USA). Biotin hydrazide, sodium cyanoborohydride (NaCNBH₄), spin desalting columns, centrifuge columns, and streptavidin agarose resin were obtained from Thermo Scientific (Waltham, MA, USA). Other chemicals were of the highest grade available. Cigarettes were purchased from a market.

RMS13 cells were obtained from the American Type Culture Collection (Manassas, VA, USA) and maintained in RPMI 1640 medium (Life Technologies, Eggenstein, Germany), supplemented with 10% fetal calf serum (FCS) (Biochrom, Berlin, Germany), 1 mM glutamine, and 1% penicillin/streptomycin (Invitrogen, Karlsruhe, Germany). PI3K/mTOR inhibitor PI103 (11 (link)) was purchased from Merck Millipore (Darmstadt, Germany), RSL3 from InterBIOScreen Ltd. (Moscow, Russia), Auranofin from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA). All chemicals were purchased from Sigma (Deisenhofen, Germany) unless indicated otherwise.

Auranofin (Santa Cruz, CA) and Piperlongumine (Sigma, St. Louis, MO) were suspended in dimethyl sulfoxide and stored in volumes of 1 mL at −20°C. Human gastric cancer cell lines SGC-7901, BGC-823 and KATO III were purchased from the Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences. The cells were routinely cultured in RPMI 1640 medium (Gibco, Eggenstein, Germany) containing 10% heat-inactivated fetal bovine serum (Gibco, Eggenstein, Germany), 100 units/mL penicillin, and 100 μg/mL streptomycin in a humidified cell incubator with an atmosphere of 5% CO₂ at 37°C. Antibodies including anti-p-PERK, anti-Bcl-2, anti-Bax, anti-cleaved PARP, anti-caspase-3 p30/17, anti-GAPDH, goat anti-mouse IgG-HRP and donkey anti-rabbit IgG-HRP were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Antibodies including anti-CHOP, anti-ATF4, anti-p-eIF2α, anti-Cleaved caspase-3 were purchased from Cell Signaling Technology (Danvers, MA). FITC Annexin V apoptosis Detection Kit I and Propidium Iodide (PI) were purchased from BD Pharmingen (Franklin Lakes, NJ).