Sensitivity of Ewing’s sarcoma family of tumors (ESFT) to fenretinide-induced cell death is dependent on sustained activation of p38MAPK but not JNK Stephen S. Myatt and Susan A. Burchill. Cancer Research UK Clinical Centre, Leeds, United Kingdom. Background: ESFT cells are more sensitive to fenretinide-induced cell death than the neuroblastoma cell line SH-SY5Y. We have previously suggested this may be effected through p38MAPK (Myatt and Burchill, Proc. Am. Assoc. Cancer Res. 2004). In the current study, we have examined the activation of the stress-activated kinases p38MAPK and JNK, and their role in fenretinide-induced cell death of ESFT cells. Methods: The effect of fenretinide on viable cell number and apoptosis was examined using the trypan blue exclusion assay and flow cytometry of annexin V-propidium iodide stained cells. The roles of p38MAPK, JNK, cytochrome c and death receptors in fenretinide-induced death were investigated by western blot and inhibitor experiments. Accumulation of ROS, and Äøm were investigated by flow cytometry. Results: Fenretinide induces cell death in all six ESFT cell lines (p<0.001) examined in a dose (0.75-3ìM) and time (<24h) dependent manner. All ESFT cell lines were more sensitive to fenretinide than the neuroblastoma cell line SH-SY5Y (p<0.001). p38MAPK was activated within 15min of fenretinide treatment in ESFT cells, and was sustained up to 8h. p38MAPK was not activated until 4h after treatment in the SH-SY5Y cells. Inhibition of p38MAPK using the selective kinase inhibitor SB202190 partially rescued fenretinide-mediated cell death (p<0.001) and PARP cleavage in ESFT but not in SH-SY5Y cells. Accumulation of ROS occurred within 5min exposure to fenretinide (1.5ìM), and was 2.4 fold higher in the ESFT compared to neuroblastoma cells. Inhibition of ROS using vitamin c (100ìM) or specific inhibitors of ROS rescued cells from cell death (p<0.001). Inhibition of ROS accumulation using vitamin c blocked activation of p38MAPK, but SB202190 had no effect on accumulation of ROS. JNK was not activated in SK-N-MC ESFT cells, but was activated 4h after treatment in TC-32 and SK-ES cells. Where activation of JNK was observed, it was dependent on ROS accumulation but not p38MAPK activity. Following 8h treatment Äøm were observed in ESFT cells, and was both ROS and p38MAPK dependent. Furthermore, release of cytochrome c was detected following 16h treatment with fenretinide (1.5ìM). Inhibition of Äøm using bongkrekic acid (100ìM) rescued 79% of cell death (1.5ìM fenretinide; 24h). Fenretinide-induced cell death appears to be associated with up-regulation of cell death receptor expression, which may be effected through sustained activation of p38MAPK. Conclusions: These data demonstrate that the high sensitivity of ESFT cells to fenretinide is in part dependent on the rapid and sustained activation of p38MAPK, which may transcriptionally regulate death receptor expression. Abstract 4545. AACR 2005 Remember we are NOT Doctors and have NO medical training. This site is like an Encylopedia - there are many pages, many links on many topics. Support our work with any size DONATION - see left side of any page - for how to donate. You can help raise awareness of CAM.