| Our recent phase II clinical trial suggests that elevation of serum C 18-ceramide might be a novel serum biomarker of chemotherapy response. Mechanistically, the pro-cell death role of C18-ceramide in cancer has not been fully elucidated, yet has an obvious clinical benefit. One novel mechanism shown here, is that bioactive C 18- ceramide selectively binds I2PP2A/SET oncoprotein within two anti-parallel beta sheet stabilized loops and an alpha helix including the K209 and Y122 residues to relieve tumor suppressor, PP2A from I2PP2A, leading to growth inhibition. I2PP2A/SET - C18- ceramide binding was dependent on the nuclear localization of I2PP2A/SET, and forcing its expression in the endoplasmic reticulum (ER) altered its selectivity for C18-ceramide. Biologically, studies revealed that ceramide mediates c-Myc degradation via its PP2A-dependent dephosphorylation at S62, and expression of c-Myc mutants with S62A or S62D conversions resulted in resistance to ceramide-mediated degradation.;Examination of a lung tissue microarray and fresh frozen lung tumors revealed that I2PP2A/SET was significantly overexpressed in lung tumor tissue when compared to adjacent non-cancerous lung tissue. Additionally, low levels of C18-ceramide were found in lung cancer tissue. Thus, the combination of high levels of I2PP2A/SET and low levels of C18-ceramide seems to promote lung tumor growth. Therapeutically, targeting I2PP2A/SET with a sphingolipid based drug, could re-constitute ceramide tumor suppressor signaling. Remarkably, the sphingolipid derivative drug, FTY720 (Fingolimod), directly bound I2PP2A/SET, which relieved and activated ceramide-PP2A tumor suppressor signaling against lung cancers in situ and in vivo . Stable knockdown of I2PP2A/SET in lung cancer cells caused a decrease in lung cancer cell growth and tumor growth, however, made these cells more resistant to FTY720 treatment. Reconstitution of I2PP2A/SET in lung cancer cells lead to the restoration of FTY720 efficacy in-situ and in-vivo. Interestingly, phosphorylation of FTY720 by sphingosine kinase-2 was not required to bind/inhibit I2PP2A/SET for inducing ceramide-PP2A dependent tumor suppression.;Lastly, FTY720's anti-cancer mechanism seems to be mediated through RIP1-dependent necroptosis. In fact, RIP1 inhibitor, necrostatin, and RIP1 siRNA or genetic loss of RIPK1 ameliorated FTY720 cell killing effect in lung cancer cells. Reconstitution of RIPK-/- MEF's with WT- or death-domain-deleted RIPK1, but not its kinase-domain-deletion mutant, restored FTY720-mediated necroptosis. |
