In this report we extend to study this combination in colon and prostate cancer cells. Both STI571 and TRAIL alone have been reported to exert antitumor activity in colon cancer cells [28,29]. Intriguingly, in this study we Volasertib leukemia found that STI571 can attenuate TRAIL-induced cytotoxicity in colon cancer cells, whereas it cannot affect TRAIL’s effect in prostate cancer cells. We presented evidence that c-Abl mediation of TRAIL-induced JNK and p38 activation is involved in the death of colon cancer cells, but not of prostate cancer cells. Moreover, p73 is the downstream effector of c-Abl which propagates signals to JNK and p38. Methods Reagents TRAIL was purchased from PeproTech (London, UK). STI571 was kindly provided by Norvartis Pharma AG (Basel, Switzerland).

Rabbit monoclonal antibodies specific for caspase 3 and 8, phosphorylated p38, JNK, ERK, and c-Abl were obtained from Cell Signaling Technology (Beverly, MA, USA). Mouse antibodies for c-Abl, JNK1, p38, and ��-actin were from Santa Cruz Biotechnology (Santa Cruz, CA, USA). The p73 antibody was purchased from BD Pharmingen Technical (San Jose, CA, USA). SB203580, SP600125, and z-Val-Ala-Asp-fluromethylketone (zVAD) were purchased from Calbiochem (San Diego, CA, USA). GST-CRK (120-225) protein was obtained from Merck Millipore (Darmstadt, Germany). All other chemicals were obtained from Sigma Aldrich (St. Louis, MO, USA). Cell culture Human colon cancer HCT116 and SW480 cells, CML K562 cells, and prostate cancer PC3 and LNCaP cells obtained from American Type Culture Collection (Manassas, VA, USA) were grown in DMEM.

All media were supplemented with 10% (v/v) heat inactivated FBS, 100 U/ml penicillin and 100 ��g/ml streptomycin. Cells were incubated at 37��C in a humidified atmosphere of 5% CO2 in air and were routinely sub-cultured every 2-3 days. Measurement of cell viability Cell viability was determined by 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide (MTT) at 1 mg/ml for 30 min, then cells were dissolved in 100% DMSO. The net absorbance (OD550 nm-OD630 nm) was determined and indicated the enzymatic activity of mitochondria and cell viability. Apoptotic assay After drug treatment, cells were harvested and washed twice with PBS and fixed in iced 70% ethanol, then stored at -20��C overnight. DNA extraction buffer (0.2 M Na2HPO4 and 0.1 M citric acid; pH 7.

8) was added at room temperature for 30 min. Cells were then incubated in PBS containing 1 mg/ml RNaseA and 40 ��g/ml propidium iodide (PI) for 30 min in the dark at room temperature. Using a FACScan flow cytometer (BD Biosciences), 104 cells were counted, and Entinostat a lower DNA content than that of the G0/G1 phase indicated apoptotic cells. Western blotting Cells were lysed by the addition of cold RIPA buffer [150 mM NaCl, 50 mM Tris HCL, 0.