Anisomycin is known to inhibit protein synthesis and induce ribotoxic stress. In this study, we investigated whether anisomycin treatment could modulate TRAIL-mediated apoptosis in human renal carcinoma Caki cells. We found that anisomycin treatment (10-15 nM) alone had no effect on the level of apoptosis, but a combination treatment of anisomycin and TRAIL significantly increased the level of apoptosis in human renal carcinoma (Caki, ACHN and A498), human glioma (U251MG), and human breast carcinoma (MDA-MB-361 and MCF7) cells. Anisomycin treatment led to the down-regulation of Bcl-2 expression at the transcriptional level, and the over-expression of Bcl-2 inhibited the apoptosis induced by the combination treatment of anisomycin and TRAIL. Furthermore, anisomycin treatment resulted in the down-regulation of c-FLIP(L) and Mcl-1 at the post-transcriptional level, and the over-expression of c-FLIP(L) and Mcl-1 blocked the induction of apoptosis caused by the combination treatment of anisomycin with TRAIL. In contrast, anisomycin treatment had no effect on the levels of TRAIL-mediated apoptosis in mouse kidney cells (TMCK-1) or normal human skin fibroblasts (HSF). Cumulatively, our study demonstrates that anisomycin treatment enhances TRAIL-mediated apoptosis through the down-regulation of Bcl-2, c-FLIP(L) and Mcl-1 at the transcriptional or post-transcriptional level.