C2-Ceramide Induces Cell Death and Protective Autophagy in Head and Neck Squamous Cell Carcinoma Cells

Ceramides are second messengers involved in several intracellular processes in cancer cells, amongst others. The aim of this study was to evaluate the anti-tumor efficacy of C2-ceramide (C2-Cer; N-acetyl-d-sphingosine) by investigating cell death and autophagy in head and neck squamous cell carcinoma (HNSCC) cells. C2-Cer showed concentration-dependent cytotoxicity in HN4 and HN30 cell lines. It simultaneously induced caspase-3-independent apoptosis and programmed necrosis. C2-Cer markedly increased the expression level of microtubule-associated protein 1 light chain 3B (LC3B) type II associated with protective autophagy. An autophagy inhibitor enhanced C2-Cer-mediated cytotoxicity, while a programmed-necrosis inhibitor produced the opposite effect. Furthermore, C2-Cer up-regulated the phosphorylation of extracellular signal-regulated kinase 1/2, but down-regulated its downstream substrate phospho-mammalian target of rapamycin (p-mTOR) during the autophagy process. These results suggested that C2-Cer exerts anti-tumor effects by inducing programmed apoptosis and necrosis in HNSCC, and these cytotoxic effects are enhanced by an autophagy inhibitor.