Human TNBC cell lines MDA-MB-231 (ATCC® CRM-HTB-26™) and HS578T (ATCC® HTB-126™) (ATCC, VA, USA) were cultured in Dulbecco’s modified eagle’s medium (DMEM, 30-2002, ATCC) at 37℃ with 10% fetal bovine serum (FBS, 16000044, GIBCO, NY, USA). In order to construct Gef-resistant cells, MDA-MB-231 and HS578T cells were cultured in the medium with increasing concentration of Gefitinib for 11 months [the highest concentration of Gefitinib was 3 μM (21 )]. Finally, the Gef-resistant cells MDA-MB-231/Gef and HS578T/Gef were stored in the medium containing 3 μM Gefitinib (22 ). Gefitinib was purchased from MedChemExpress (HY-50895, Shanghai, China).
The Gef-resistant cells were assigned into sh-NC group (treated with lentivirus small hairpin RNA-negative control), sh-GPX4 group (treated with lentivirus sh-GPX4), sh-NC + DMSO group (treated with sh-NC + dimethyl sulphoxide), sh-GPX4 + DMSO (treated with lentivirus sh-GPX4 + DMSO), and sh-GPX4 + Ferrostatin-1 group (treated with lentivirus sh-GPX4 and 1 μM Ferrostatin-1) (23 (link)). Ferrostatin-1 is an inhibitor of ferroptosis and purchased from MedChemExpress (HY-100579). The titer of lentivirus was 1× 108 Tu/ml.
The Gef-resistant cells were assigned into sh-NC group (treated with lentivirus small hairpin RNA-negative control), sh-GPX4 group (treated with lentivirus sh-GPX4), sh-NC + DMSO group (treated with sh-NC + dimethyl sulphoxide), sh-GPX4 + DMSO (treated with lentivirus sh-GPX4 + DMSO), and sh-GPX4 + Ferrostatin-1 group (treated with lentivirus sh-GPX4 and 1 μM Ferrostatin-1) (23 (link)). Ferrostatin-1 is an inhibitor of ferroptosis and purchased from MedChemExpress (HY-100579). The titer of lentivirus was 1× 108 Tu/ml.
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