To determine whether the apoptotic machinery of thyroid cancer cells is functional and could be activated for tumoricidal purposes, we examined the apoptosis induced by the cytokines TNF-α, Fas and TRAIL in thyroid cancer cell lines, NPA and SW579. Interestingly, out of these cytokines, only TRAIL was able to trigger significant apoptosis. The tumoricidal effect of TRAIL was further enhanced by CHX, suggesting the presence of CHX-sensitive inhibitor (s) of apoptosis in these thyroid cancer cell lines. The anti-apoptotic proteins like FLAME-1, Bcl-2 and Bcl-xL are believed to be such CHX-sensitive inhibitors in various types of cancer cells. We, however, provide the evidence using NPA and SW579 cell lines that these proteins were not affected by the CHX treatment in thyroid cancer cells. The apoptosis of thyroid cancer cells was mediated by the classical activation of caspases that in turn activated the DNA Fragmentation Factor (DFF-45). To elucidate the role of individual caspases in TRAIL-mediated apoptosis, the inhibitory effects of several general and specific tetra-peptide caspase inhibitors were studied. The inhibitors of caspase-1,-6,-8, and-9 as well as general upstream inhibitors of apoptosis could dramatically inhibit TRAIL-induced apoptosis in thyroid cancer cells. Caspase-2 and-3 inhibitors, on the other hand, had no significant effect. When the cells were treated with either agonistic Fas antibody (CH11) or TNF-α, no apoptotic changes were observed. The apoptosis induced by agonistic Fas Ab could be seen only after a prolonged exposure (24 h) to CHX, whereas TNF-α had no effect even in the presence of CHX. The efficacy of TRAIL was also tested on other types of thyroid cancer cells like ARO, FRO (anaplastic carcinoma) and TPC-1 (papillary carcinoma) and compared to that triggered by other death inducing cytokines FasL and TNF-α. Again TRAIL was more potent in triggering apoptosis than Fas and TNF-α. Since TRAIL is effective in selectively killing thyroid tumor cells without affecting normal thyrocytes and also does not cause organ toxicity and inflammation in vivo, its potential for the treatment of thyroid cancer seems very promising.