The aim of this study was to understand the molecular requirements for cisplatin-induced apoptosis in human head and neck squamous cell carcinoma (HNSCC) cell death. Cisplatin induced apoptosis in HNSCC cell lines, HSC-2, HSC-3 and HSC-4 in a dose-dependent manner. However, cisplatin did not induce the expression of Fas-ligand mRNA or upregulation of Fas protein. By caspase activation assays, cisplatin induced Caspase-3 (Casp-3), -8 and -9 activation. In all three lines tested, the use of a specific inhibitor of Casp-9 almost completely blocked cisplatin-induced apoptosis, while the use of Casp-3 and -8 inhibitors resulted in a partial blockade of cisplatin-induced apoptosis. Our results strongly suggest that Casp-9-dependent apoptosis plays an important role in cisplatin-induced HNSCC apoptosis.