Gsk1120212

NVP-BEZ235 (BEZ235), NVP-BKM120 (Buparlisib, BKM120), GSK1120212 (Trametinib, JTP-74057), PD0325901, and AZD6244 were purchased from Chemie Tek (Indianapolis, IN). Temozolomide, doxorubicin and paclitaxel were purchased from Sigma. For in vivo studies, BEZ235 and BKM120 were resuspended in NMP (N-methyl-2-pyrrolidone):PEG300 1:9 v:v, and GSK1120212 and PD0325901 were resuspended in 0.5% hydroxypropyl-methylcellulose (Sigma):0.2% Tween 80 (Sigma) (Gilmartin et al., 2011 (link); Kinross et al., 2011 (link)). BKM120, GSK1120212 and PD0325901 were given once daily by oral gavage at 10 ml/kg. Mice were treated with BEZ235 on a 5 days on, 2 days off schedule (Liu et al., 2009 (link)). Vehicle- and drug-treated mice were closely monitored on a daily basis for clinical signs as described above.

We maintained human serous ovarian cancer cell lines including OVCA420, OVCA433, OVCA429, TYK-nu, SKOV8, CAOV3, DOV13, HEYA8, A2780 and SKOV3 [8 (link)], in RPMI 1640 (Nacalai Tesque, Kyoto, Japan) supplemented with 10% fetal bovine serum (FBS) and 100 IU/ml penicillin and 100 μg/ml streptomycin (Nacali Tesque). The immortalized human ovarian surface epithelial cell line, HOSE-E7 [20 (link)], kindly provided by Dr. Katabuchi at Kumamoto University, was cultured in DMEM/F12 (Thermo Fisher Scientific, Waltham, USA) supplemented with 10% FBS and 100 IU/ml penicillin and 100 μg/ml streptomycin (Nacalai Tesque). 293FT cells were purchased from Thermo Fisher Scientific, and cultured in DMEM (Thermo Fisher Scientific) supplemented with 10% FBS and 100 IU/ml penicillin and 100 μg/ml streptomycin. All cells were seeded into Cellstars® tissue culture plates (Greiner, Frickenhausen, Germany) in a humidified incubator containing 5% CO₂ at 37°C. All cell lines were regularly tested for mycoplasma.
As an inhibitor of the p38 MAPK pathway, we used SB203580 (Sigma-Aldrich, St.Louis, USA). To inhibit the ERK1/2 pathway, we used GSK1120212 (Sigma-Aldrich).