Abstract
BackgroundCeramide is a bioeffector that mediates various cellular processes, including apoptosis. However, the mechanism underlying ceramide function in apoptosis is apparently cell type-dependent and is not well-understood. We aimed at identifying molecular targets of ceramide in metastatic human colon and breast cancer cells, and determining the efficacy of ceramide analog in suppression of colon and breast cancer metastasis.MethodsThe activity of and mechanism underlying ceramide as a cytotoxic agent, and as a sensitizer for Fas-mediated apoptosis was analyzed in human cell lines established from primary or metastatic colon and breast cancers. The efficacy of ceramide analog LCL85 in suppression of metastasis was examined in preclinical mouse tumor models.ResultsExposure of human colon carcinoma cells to ceramide analog LCL85 results in apoptosis in a dose-dependent manner. Interestingly, a sublethal dose of LCL85 increased C16 ceramide content and overcame tumor cell resistance to Fas-mediated apoptosis. Subsequently, treatment of tumor cells with exogenous C16 ceramide resulted in increased tumor cell sensitivity to Fas-mediated apoptosis. LCL85 resembles Smac mimetic BV6 in sensitization of colon carcinoma cells to Fas-mediated apoptosis by inducing proteasomal degradation of cIAP1 and xIAP proteins. LCL85 also decreased xIAP1 and cIAP1 protein levels and sensitized metastatic human breast cancer cells to Fas-mediated apoptosis. Silencing xIAP and cIAP1 with specific siRNAs significantly increased the metastatic human colon carcinoma cell sensitivity to Fas-mediated apoptosis, suggesting that IAP proteins mediate apoptosis resistance in metastatic human colon carcinoma cells and ceramide induces IAP protein degradation to sensitize the tumor cells to apoptosis induction. Consistent with its apoptosis sensitization activity, subtoxic doses of LCL85 suppressed colon carcinoma cell metastatic potential in an experimental lung metastasis mouse model, as well as breast cancer growth and spontaneous lung metastasis in an orthotopic breast cancer mouse model.ConclusionWe have identified xIAP and cIAP1 as molecular targets of ceramide and determined that ceramide analog LCL85 is an effective sensitizer in overcoming resistance of human cell lines established from metastatic colon and breast cancers to apoptosis induction to suppress metastasis in vivo.
Highlights
Ceramide is a bioeffector that mediates various cellular processes, including apoptosis
We demonstrated that aromatic ceramide analog LCL85 effectively overcomes metastatic human colon and breast cancer cell resistance to Fas-mediated apoptosis at least partially through inducing proteasomal degradation of cIAP1 and xIAP in vitro
Ceramide analog effectively sensitizes metastatic human colon and breast cancer cell apoptosis resistance Ceramide analogs of B13 and LCL85 were tested for their cytotoxicity against human colon carcinoma cell lines (6 primary and 5 metastatic human colon cancer cell lines)
Summary
Ceramide is a bioeffector that mediates various cellular processes, including apoptosis. Compelling experimental data from both human cancer patients and mouse tumor models indicate that the Fas-mediated apoptosis pathway plays a key role in suppression of cancer development and in host cancer immunosurveillance [1,2,3]. Human cancer genomics data indicate that Fas is not significantly focally amplified across a dataset of 3131 tumors, but is significantly focally deleted across the entire dataset of these 3131 tumors, including human colorectal cancer These data strongly suggest that Fas functions as a tumor suppressor
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