The amplified TRAIL induced caspase activation by overexpressed c Myc counteract the anti apoptotic activity of DNA PKcs by increasing proteolytic cleavage in the metastatic cancer cells It has been known that a constitutively active Akt is an important Tofacitinib baldness regulator of TRAIL sensitivity, and inhibition of Akt activation by its pharmacological inhi bitor or knockdown Inhibitors,Modulators,Libraries of its expression by siRNA sensi tizes TRAIL resistant cells to TRAIL. The activation of Akt Inhibitors,Modulators,Libraries by phosphorylation of Ser 473 is mediated by DNA PKcs. However, the metastatic cancer cells were sensitive to TRAIL, despite that the metastatic cells have higher level of DNA PKcs com pared with their primary cells, as shown above. There fore, we determined the levels of DNA PKcs and pAkt in the metastatic cells after treatment with TRAIL.

In metastatic PC3 MM2 and KM12L4A cells, DNA PKcs was cleaved and consequently the level of DNA PKcs was decreased after exposure to TRAIL and this result was accompanied with decrease of pAkt level, whereas the levels of DNA PKcs and pAkt were maintained in the primary PC3 and KM12 cells after Inhibitors,Modulators,Libraries exposure to TRAIL. Since c Myc could control the TRAIL sensitivity, and DNA PKcs is a substrate of caspase 3, we deter mined whether depletion of c Myc could block degrada tion of DNA PKcs. When PC3 MM2 cells were transfected with c Myc siRNA, TRAIL induced cleavage of DNA PKcs was reduced compared with transfection with scrambled siRNA. These results were followed by prevention of cleavage of DNA PKcs as well as PARP in the TRAIL treated cells by pretreatment with Z DEVD FMK, a caspase 3 specific inhibitor.

Inhibitors,Modulators,Libraries These Inhibitors,Modulators,Libraries results suggest that the amplified TRAIL induced cas pase activation by over expressed c Myc may curtail the anti apoptotic activity of DNA PKcs by increasing its proteolytic cleavage in the metastatic cancer cells. Suppression of DNA PKcs is associated with hypersensitivity to TRAIL induced cytotoxicity To investigate the direct role of DNA PKcs in the sus ceptibility of the metastatic cells to TRAIL, we used siRNA to knockdown DNA PKcs expression and deter mined its effect on TRAIL sensitivity. After transfection of PC3 or KM12 cells with siRNA against DNA PKcs or scrambled siRNA, the expression of DNA PKcs was effi ciently suppressed by DNA PKcs siRNA as compared to the control cells and its expression was further decreased by TRAIL treatment.

This result was followed by the hypersensitivity to TRAIL induced reduction of DNA PKcs/pAkt levels, activation of cas pases, PARP cleavage, and up regulation of Bax in PC3 and KM12 cells after transfection those with DNA PKcs siRNA as compared to the cells transfected with scrambled siRNA. Furthermore, the knockdown of DNA PKcs with specific siRNA significantly increased TRAIL induced apoptosis in PC3 and KM12 cells.