Bms 911543

The anti-PD-L1 mAb (clone 405.9A11) was previously validated (26 ) by Dr. Gordon J. Freeman (Dana-Farber Cancer Institute, Boston, MA), anti-pJAK2(Y1007 and Y1008) (Abcam, Cambridge, UK) were used for IHC staining. The anti-PD-L1-PE (BD Pharmingen, San Jose, CA) anti-HLA-ABC-FITC mAb (clone w6/32, E-biosciences, San Diego, CA), IgG1-PE isotype control, pSTAT1 Tyr701-PE, STAT1-PE and STAT3-PE (BD Biosciences, San Jose, CA), phospho-AKT(Thre308)-PE and primary anti-p44/42 MAPK(Erk1/2) and phospho-p44/42 MAPK(Erk1/2) (Thr202/Tyr204) and secondary anti-rabbit-PE antibodies (Cell Signaling, Danvers, MA). WB antibodies included rabbit anti-human JAK2, pJAK2 (y1007/1008), total AKT, pAKT, pERK and mouse anti-human β-actin (Cell signaling, Danvers, MA). IFNγ (R&D systems Minneapolis, MN) was used at 10IU/mL. IFNα2a (PBL Interferon Source, Piscataway, NJ) was used at 1000 IU/mL. JAK2 inhibitor BMS-911543 was characterized previously (25 (link)), provided by Bristol-Myers Squibb and used at 10uM. JAK1/3 inhibitor (ZM39923) (Tocris bioscience, Bristol, United Kingdom) was characterized previously (27 (link), 28 (link)) and was used at 10uM. Wortmannin (Cell Signaling, Danvers, MA) was used at a 1uM. PI3Kα110 subunit inhibitor (BYL-719) was used at a 1uM and MEK1/2 inhibitor (PD0325901) was used at 5uM (Tocris Bioscience, Bristol, UK).