Signaling pathways mediating the potentiation of TRAIL-induced cell death by HGF. A. DAOY cells were pre-incubated with or without inhibitors for 30 min, and then treated with HGF for 72 h followed by TRAIL for 24 h. Cell viability was analyzed by MTT assay. The c-Met inhibitor PHA-665752 (100 nM) significantly reduced the cell death promoting effect of HGF. The PI3 kinase inhibitor LY294002 (30 μM) or wortmannin (1 μM) slightly increased the cell death rate induced by TRAIL + HGF. The ERK inhibitor PD98059 (30 μM) as well as the JNK pathway inhibitor CEP-11004 (10 μM) had no effect on the cell death induced by TRAIL + HGF. B. DAOY cells were pre-incubated with c-Met inhibitor PHA-665752 (100 nM) for 30 min before adding HGF. Phosphorylation of c-Met in cells treated with HGF or HGF+TRAIL was inhibited by PHA-665752. C. Western blot analysis with anti-JNK and anti-phospho-JNK (Ser 198) antibodies was used to determine JNK activation. In DAOY cells, HGF induced JNK activation as early as 5 min and lasted for 30 min. C-Jun was also activated within 10 min of HGF treatment. Data represent mean ± SE (N = 9, * P < 0.001).