Flo 1

The human distal oesophageal adenocarcinoma cell lines, FLO-1 and KYAE-1, were purchased from European Collection of Cell Cultures (Salisbury, UK). The human pancreatic cancer cell line AsPc-1 was purchased from the American Type Culture Collection (Manassas, VA, USA). AsPc-1^shTAK1 cells stably expressing small hairpin RNA sequences to knockdown the expression of TAK1 has been previously described in Melisi et al (2011) (link). Distal oesophageal adenocarcinoma and pancreatic cancer cell lines were maintained in RPMI 1640 and Dulbecco's modified Eagle's media (LONZA, Basel, Switzerland), respectively, supplemented with 10% heat-inactivated FBS, 20 mmol l⁻¹ HEPES (pH 7.4), penicillin (100 UI ml⁻¹), streptomycin (100 mg ml⁻¹), and 4 mmol l⁻¹ glutamine (ICN Biomedicals Ltd., Irvine, CA, USA) in a humidified atmosphere of 95% air and 5% CO₂ at 37 °C. The TAK1 kinase activity was targeted using (5Z)-7-oxozeaenol TAK1 kinase selective inhibitor (TOCRIS bioscience, Bristol, UK). For in vitro assays, (5Z)-7-oxozeaenol was dissolved in 100% dimethyl sulfoxide (DMSO) at a stock concentration of 10 mM. Cell irradiation was performed using a GammaCell 40 irradiator (Best Theratronics Ltd., Ottawa, Canada) as previously described in Melisi et al, 2009 (link).