Bisindolylmaleimide IX facilitates extrinsic and initiates intrinsic apoptosis in TNF-alpha-resistant human colon adenocarcinoma COLO 205 cells

Human COLO 205 colon adenocarcinoma cells are immune to extrinsic Apoptosis induced by immunomodulatory cytokines. Among the antiapoptotic mechanisms responsible for the immune escape, the overexpression of the cFLIP protein seems to be critical. cFLIP appears to inhibit the TNF-alpha-induced death receptor signal. The application of the metabolic inhibitor bisindolylmaleimide IX (Bis-IX), known as a potent PKC repressor, sensitized COLO 205 cells to TNF-alpha-mediated Apoptosis. The Western-blot analysis revealed that the susceptibility of human COLO 205 cells to apoptogenic stimuli resulted from time-dependent reduction in cFLIP(L) and TRADD protein levels. At the same time, the level of FADD protein was up-regulated. Additionally, the combined TNF-alpha and Bis-IX treatment caused cleavages of Bid and procaspase-9, as well as cytochrome c release. Thus, the evidence of this study indicates that Bis-IX facilitates the death receptor signal mediated by TNF-R1. Moreover, Bis-IX alone initiated intrinsic Apoptosis, which could be abolished by Bcl-2 delivery. It heralds the involvement of mitochondria in caspase-8-independent intrinsic Apoptosis. In turn, the treatment with bisindolylmaleimide III (Bis-III) did not assist TNF-alpha-dependent Apoptosis.