Upci scc154

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.

HNSCC cell lines Cal 27, Cal 33, UDSCC 2, UPCISCC 040, UPCISCC 099, UPCISCC 131, UPCISCC 154 were purchased from ATCC (Manassas, VA, USA), DSMZ (Braunschweig, Germany), or obtained from Prof. Thomas Hoffmann (University of Ulm, Germany) and authenticated after retrieval by short tandem repeat (STR) typing (service by DSMZ) [45 (link)–47 (link)]. Cell lines were cultured in DMEM supplemented with 10% FCS, 100 U/ml penicillin, and 100 µg/ml streptomycin (all from Thermo Scientific, Schwerte, Germany) at 37 °C and 7.5% CO₂. Human hTERT-immortalized keratinocytes OKF 6 were cultured in a 2 + 1 + 1 Keratinocyte-SFM/DMEM/F12 nutrient mixture, supplemented with 0.3 ng/ml rEGF, 35 µg/ml bovine pituitary extract, 155 µM CaCl₂, 1.5 mM Glutamax, 25 U/ml penicillin, and 25 µg/ml streptomycin (Thermo Scientific) [48 (link)]. All cell lines were routinely tested negative for mycoplasma contamination. Cells were irradiated at the indicated doses using an RS225 X-ray cabinet (X-Strahl, Camberley, UK) operated at 200 kV/10 mA (Thoraeus filter, 1 Gy in 63 s).

A431 (CRL-1555; RRID:CVCL_0037), CAL27 (CRL-2095; RRID:CVCL_1107), CaSki (CRL-1550; RRID:CVCL_1100), H23 (CRL-5800; RRID:CVCL_1547), Calu6 (HTB-56; RRID:CVCL_0236), and UPCI:SCC154 (CRL-3241; RRID:CVCL_2230) cell lines were purchased from ATCC (Manassas, VA, USA). A431, CAL27, and Calu6 were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM, #30-2002). CaSki and H23 were cultured in RPMI 1640 (#30-2001). UPCI:SCC154 was cultured in Eagle’s Minimal Essential Medium (EMEM, #30-2003) supplemented with 2 mM L-glutamine. Cell culture medium was obtained from ATCC (Manassas, VA, USA). Cells were supplemented with 10% fetal bovine serum (FBS, #12484028) and 5% penicillin-streptomycin (#15140122) and maintained at 37 °C with 5% CO₂ [30 (link),31 (link)]. Cells were periodically evaluated for mycoplasma contamination by DAPI stain for extra-nuclear DNA and MycoFluor^TM Mycoplasma detection kit (#M7006). Reagents listed were obtained from Thermo Fisher Scientific (Waltham, MA, USA). Human subjects in this study were patients with histopathologically-verified SCCs. This study was approved by Institutional Review Board, The Ottawa Hospital (IRB Protocol # 20150896-01H) and Research Institute of the McGill University Health Centre (IRB Protocols # 2018-4128 and 2022-8414).