C6-ceramide synergistically potentiates the anti-tumor effects of histone deacetylase inhibitors via AKT dephosphorylation and α-tubulin hyperacetylation both in vitro and in vivo

Abstract

Histone deacetylase inhibitors (HDACIs) have shown promising anti-tumor effects for a variety of malignancies, however, many tumors are reportedly resistant to them. In this study, we made a novel discovery that co-administration of HDACIs (Trichostatin A (TSA) and others) and exogenous cell-permeable short-chain ceramide (C6) results in striking increase in cancer cell death and apoptosis in multiple cancer cells. These events are associated with perturbations in diverse cell signaling pathways, including inactivation of Akt/mTOR and increase in α-tubulin acetylation (both in vivo and in vitro). TSA interacts in a highly synergistic manner with C6-ceramide to disrupt HDAC6/protein phosphatase 1 (PP1)/tubulin complex, to induce α-tubulin hyperacetylation, and to release and activate PP1, which then leads to AKT dephosphorylation and eventually causes cancer cell death. Interestingly, TSA itself results in short-term ceramide accumulation, which as a result of metabolic (glycosylation) removal, does not result in evident increase of cancer cell death. However, adding C6-ceramide led to a very pronounced increase in ceramide level and marked increase in cell death. Importantly, the effective synergistic anti-tumor activity of TSA plus C6-ceramide is also seen in in vivo mice xenograft pancreatic and ovarian cancer models, indicating that this regimen (HDACI plus C6-ceramide) may represent a more effective form of therapy against pancreatic and ovarian carcinoma.

Sphingosine kinase‐1 inhibition sensitizes curcumin‐induced growth inhibition and apoptosis in ovarian cancer cells

“…Consistent with previous findings, (10,21) we observed a moderate ceramide accumulation after 24 h of curcumin (25 lM) treatment in CaOV3 cells (Fig. 5A).…”

“…(3,4,33) Recent studies suggest that the phosphatase-dependent mechanism might be responsible for Akt inhibition by curcumin. (33) Previous studies, including ours, (21) have indentified ceramide-activated protein phosphatase (CAPP) as ceramide regulated enzymes. CAPP includes serine/threonine phosphatases 2A (PP2A) (34)(35)(36) and protein phophatase-1 (PP1).…”

“…( [21][22] Generation of a CaOV3 line stably expressing a dominant-negative p38 mutant. The full length of p38 was cloned into pCB7 vector as a HindIIII fragment.…”

“…As previously reported, (21) a plasmid encoding a constitutively active Akt1 cDNA (Plasmid 16244) was obtained from Addgene (Cambridge, MA, USA). For transfection experiments, 4.0 lL PLUS Reagent (Invitrogen, Carlsbad, CA, USA) was diluted in 90 lL of RNA dilution water (Santa Cruz Biotechnology) for 5 min at room temperature.…”

Diverting CERT-mediated ceramide transport to mitochondria triggers Bax-dependent apoptosis

“…Some reports indicate that ceramides can promote apoptotic cell death by inhibiting phosphoinositide-3 kinase (PI3K) and Akt/PKB signaling, resulting in dephosphorylation and subsequent activation of the pro-apoptotic Bcl-2-family protein Bad (Bourbon et al, 2002;Zhu et al, 2011). Short-chain ceramides can activate protein phosphatase 2A (PP2A), which dephosphorylates and inactivates the anti-apoptotic protein BCL2 (Dobrowsky et al, 1993;Mukhopadhyay et al, 2009).…”