All cell lines were maintained in a humidified incubator at 37 °C with 5% CO2. Cell line identities were validated by short tandem repeat analysis (LabCorp, Genetica Cell Line Testing), and cultures were regularly tested for mycoplasma contamination (Lonza). The UPCI:SCC090 (CRL-3239), UPCI:SCC152 (CRL-3240), and UPCI:SCC154 (CRL-3241) cell lines were purchased from ATCC and cultured in Eagle's Minimum Essential Media (Corning) supplemented with 10% fetal bovine serum (Sigma), 1% penicillin–streptomycin (Corning), and 2 mmol/L ʟ-glutamine (GIBCO). HEK293T cells (CRL-11268) were purchased from ATCC and cultured in DMEM (Corning) supplemented with 10% fetal bovine serum and 1% penicillin–streptomycin.
Recombinant lentivirus was produced in HEK293T cells using PEI-based transfection. Briefly, psPAX2 packaging (Addgene #12260), VSV-G envelope (Addgene #12259), and UBC-driven NRF2 E79Q vectors were combined with PEI at a 3:1 ratio (μl PEI: μg DNA). Supernatants containing virus were filtered and added to SCC90, SCC152, and SCC154 cells. Transduced cells were selected with 50 μg/ml, 50 μg/ml, and 250 μg/ml hygromycin, respectively.
Recombinant lentivirus was produced in HEK293T cells using PEI-based transfection. Briefly, psPAX2 packaging (Addgene #12260), VSV-G envelope (Addgene #12259), and UBC-driven NRF2 E79Q vectors were combined with PEI at a 3:1 ratio (μl PEI: μg DNA). Supernatants containing virus were filtered and added to SCC90, SCC152, and SCC154 cells. Transduced cells were selected with 50 μg/ml, 50 μg/ml, and 250 μg/ml hygromycin, respectively.