Abstract
Hormonal therapies, including ovarian ablation, ER antagonists, and aromatase inhibitors, are the standards of care for treatment of ERα positive breast cancer. However, development of ...resistance to hormone therapies in advanced breast cancer is a major obstacle. Moreover, estrogen receptor-α (ERα)-negative breast cancer is clinically aggressive and does not respond to conventional estrogen targeted therapies. Strategies that lead to re-expression of ERα could sensitize breast cancers to selective ER modulators. FTY720/Fingolimod, a sphingosine analog, is an FDA-approved pro-drug for treatment of multiple sclerosis that also has anticancer actions that are not yet well understood. We have now found that FTY720 is phosphorylated in breast cancer cells by nuclear sphingosine kinase 2 and accumulates there. Nuclear FTY720-P in turn is a potent inhibitor of class I histone deacetylases (HDACs) that enhances histone acetylations and gene expression independently of its known effects on canonical signaling through sphingosine-1-phosphate receptors. In ERα negative human and murine breast cancer cells, FTY720 reactivated expression of silenced ERα and sensitized them to tamoxifen. Moreover, oral administration of clinically relevant doses of FTY720 to mice bearing ERα negative syngeneic breast tumors also re-expressed ERα and increased therapeutic sensitivity to tamoxifen in vivo more potently than a known HDAC inhibitor. Our work suggests that FTY720 is a promising strategy for effective treatment of conventional hormonal therapy-resistant breast cancer and triple-negative breast cancer. Supported by NIH grant RO1CA061774 and the Department of Defense BCRP program award W81XWH-14-1-0086 (S. Spiegel).
Citation Format: Nitai C. Hait, Dorit Avni, Akimitsu Yamada, Sheldon Milstien, Kazuaki Takabe, Sarah Spiegel. FTY720-P is a potent inhibitor of class I histone deacetylases that enhances histone acetylation, reactivates ERα expression, and increases hormonal therapeutic sensitivity of breast cancer. abstract. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 112. doi:10.1158/1538-7445.AM2015-112
Abstract only
Non‐alcoholic fatty liver disease (NAFLD) is a metabolic disease characterized by a spectrum of related liver disorders. Liver steatosis is an early initiator of NAFLD followed by the ...development of nonalcoholic steatohepatitis. Approximately 30% of adults in the United States are affected by NAFLD and although obesity and insulin resistance are risk factors for NAFLD, there is currently no approved therapy for this disease. To make matters worse, NAFLD can progress to liver cirrhosis and ultimately hepatocellular carcinoma. Sphingolipid metabolites, particularly ceramide and sphingosine‐1‐phosphate (S1P), have recently received attention for their potential roles in insulin resistance and hepatic steatosis. FTY720/Fingolimod, an FDA approved pro‐drug for treatment of multiple sclerosis, is phosphorylated in vivo to its active phosphorylated form by sphingosine kinase 2 and has been shown to interfere with the actions of S1P and inhibits several sphingolipid metabolic enzymes. Therefore, in this study we investigated its effect in the C57BL/6 x 129S murine model of NAFLD. Mice were fed a high fat diet (42% kcal from fat) with sugar water (HFD+SW) for 16 weeks with concurrent oral administration of a clinically relevant dose of FTY720 (0.3 mg/kg). FTY720 significantly decreased liver lipid accumulation and steatosis. Mass spectrometry analysis confirmed that FTY720 was phosphorylated to its active form, FTY720‐phosphate. Although levels of S1P and monohexosylceramide were significantly decreased in blood by administration of FTY720, no major changes in ceramide were detected. Moreover, liver sphingolipid levels including ceramide, monohexosylceramide, sphingomyelin, dihydroceramide, dihydromonohexosylceramide, and dihydrosphingomyelin were all decreased (p < 0.05) by FTY720 treatment. Additionally, FTY720 treatment caused a significant decrease in activation of Akt known to play a critical role in steatosis. Taken together, these findings support the notion that targeting sphingolipid metabolism could lead to reduced hepatic steatosis.
Support or Funding Information
Supported by NIH T32 Training Grant 5T32DK007150‐39.
The synthetic phospho-ceramide analogue-1 (PCERA-1) down-regulates production of the pro-inflammatory cytokine tumour necrosis factor-α (TNF-α) and up-regulates production of the anti-inflammatory ...cytokine interleukin-10 (IL-10) in lipopolysaccharide (LPS) -stimulated macrophages. We have previously reported that PCERA-1 increases cyclic adenosine monophosphate (cAMP) levels. The objective of this study was to delineate the signalling pathway leading from PCERA-1 via cAMP to modulation of TNF-α and IL-10 production. We show here that PCERA-1 elevates intra-cellular cAMP level in a guanosine triphosphate-dependent manner in RAW264.7 macrophages. The cell-permeable dibutyryl cAMP was able to mimic the effects of PCERA-1 on cytokine production, whereas 8-chloro-phenylthio-methyladenosine-cAMP, which specifically activates the exchange protein directly activated by cAMP (EPAC) but not protein kinase A (PKA), failed to mimic PCERA-1 activities. Consistently, the PKA inhibitor H89 efficiently blocked PCERA-1-driven cytokine modulation as well as PCERA-1-stimulated phosphorylation of cAMP response element binding protein (CREB) on Ser-133. Finally, PCERA-1 activated cAMP-responsive transcription of a luciferase reporter, in synergism with the phosphodiesterase (PDE)-4 inhibitor rolipram. Our results suggest that PCERA-1 activates a Gs protein-coupled receptor, leading to elevation of cAMP, which acts via the PKA-CREB pathway to promote TNF-α suppression and IL-10 induction in LPS-stimulated macrophages. Identification of the PCERA-1 receptor is expected to set up a new target for development of novel anti-inflammatory drugs.
Highlights • C1P and the synthetic analog PCERA-1 affect macrophages via distinct receptors. • TNFα secretion is inhibited at different stages by exogenous C1P and PCERA-1. • Exogenous C1P inhibits ...TNFα secretion at the level of TACE activity. • PCERA-1 reciprocally modulates TNFα and IL-10 transcription via CREB.
The tumor necrosis factor (TNF) receptor family member CD40 plays an essential role in the activation of antigen‐presenting cells, B cell maturation, and immunoglobulin (Ig) class switching critical ...for adaptive immunity. Although the bioactive sphingolipid metabolite sphingosine‐1‐phosphate (S1P) and the kinase that produces it, sphingosine kinase 1 (SphK1), have long been implicated in the actions of TNF mediated by engagement of TNFR1, nothing is yet known of their role in CD40‐mediated events. We have now found that ligation of CD40 activates and translocates SphK1 to the plasma membrane, leading to generation of S1P. SphK1 inhibition in human tonsil B cells, as well as inhibition or deletion of SphK1 in mouse splenic B cells, significantly reduced CD40‐mediated Ig class switching and plasma cell differentiation ex vivo. Optimal activation of downstream CD40 signaling pathways, including NF‐κB, p38, and JNK, also required SphK1. In mice treated with a SphK1 inhibitor or in SphK1–/– mice, isotype switching to antigen‐specific IgE was decreased in vivo by 70 and 55%, respectively. Our results indicate that SphK1 is important for CD40‐mediated B cell activation and regulation of humoral responses and suggest that targeting SphK1 might be a useful therapeutic approach to control antigen‐specific IgE production.—Kim, E. Y., Sturgill, J. L., Hait, N. C., Avni, D., Valencia, E. C., Maceyka, M., Lima, S., Allegood, J., Huang, W.‐C., Zhang, S., Milstien, S., Conrad, D., Spiegel, S., Role of sphingosine kinase 1 and sphingosine‐1‐phosphate in CD40 signaling and IgE class switching. FASEB J. 28, 4347–4358 (2014). www.fasebj.org
Expression of the anti-inflammatory cytokine IL-10 can be induced either by TLR agonists such as lipopolysaccharide (LPS), or by various endogenous stimuli, in particular those acting via a ...cAMP-dependent signaling pathway. We have previously reported that the synthetic phospho-ceramide analogue-1 (PCERA-1) increases cAMP level and subsequently down-regulates production of TNFalpha and up-regulates production of IL-10 in LPS-stimulated macrophages. The objective of this study was to determine the mechanism of activity of PCERA-1 and the role of cAMP in LPS-induced IL-10 production. We show here that PCERA-1 induces IL-10 production in synergism with various TLR agonists in mouse RAW264.7 macrophages. Cooperativity is evident both at the mRNA and protein levels. IL-10 production by LPS and PCERA-1 is mediated by the cAMP pathway and by the p38 MAP kinase. Phosphorylation of p38 is cooperatively accomplished by LPS and PCERA-1 or other cAMP inducers. Furthermore, the activity of PCERA-1 can be partially mimicked by a cell-permeable analog of cAMP, and blocked by the protein kinase A (PKA) inhibitor H89. Finally, in the absence of PCERA-1, the residual IL-10 induction by LPS depends on the basal cAMP level as it can be largely elevated by the phosphodiesterase (PDE)-4 inhibitor rolipram. Our results thus indicate that IL-10 induction by LPS critically depends on basal cAMP level, and that a co-stimulus by a TLR agonist and a cAMP-elevating agent results in synergistic PKA-dependent and p38-dependent IL-10 production.
Beekeepers may feed colonies with pollen supplement patties during times of pollen shortage. We tested whether the surface area of pollen patties fed to honey bee colonies affects their consumption, ...brood production and honey yields. We compared three pollen-patty sizes, of equal weight, and found that consumption increased with surface area. Consequently, brood production tended to increase with pollen-patty size, and colonies fed the patty with the largest surface area produced significantly more brood than those fed a carbohydrate-only control patty. The colonies in the large surface area treatment also tended to produce more honey, though the differences were not statistically significant, probably due to the time gap between the pollen deficiency period and the time honey was harvested. We conclude that pollen patties with greater surface area are more readily consumed by honey bee colonies and may contribute more to colony development and subsequent honey yields.
Summary The synthetic phospho-ceramide analogue-1 (PCERA-1) down-regulates production of the pro-inflammatory cytokine tumour necrosis factor-alpha (TNF-alpha) and up-regulates production of the ...anti-inflammatory cytokine interleukin-10 (IL-10) in lipopolysaccharide (LPS) -stimulated macrophages. We have previously reported that PCERA-1 increases cyclic adenosine monophosphate (cAMP) levels. The objective of this study was to delineate the signalling pathway leading from PCERA-1 via cAMP to modulation of TNF-alpha and IL-10 production. We show here that PCERA-1 elevates intra-cellular cAMP level in a guanosine triphosphate-dependent manner in RAW264.7 macrophages. The cell-permeable dibutyryl cAMP was able to mimic the effects of PCERA-1 on cytokine production, whereas 8-chloro-phenylthio-methyladenosine-cAMP, which specifically activates the exchange protein directly activated by cAMP (EPAC) but not protein kinase A (PKA), failed to mimic PCERA-1 activities. Consistently, the PKA inhibitor H89 efficiently blocked PCERA-1-driven cytokine modulation as well as PCERA-1-stimulated phosphorylation of cAMP response element binding protein (CREB) on Ser-133. Finally, PCERA-1 activated cAMP-responsive transcription of a luciferase reporter, in synergism with the phosphodiesterase (PDE)-4 inhibitor rolipram. Our results suggest that PCERA-1 activates a Gs protein-coupled receptor, leading to elevation of cAMP, which acts via the PKA-CREB pathway to promote TNF-alpha suppression and IL-10 induction in LPS-stimulated macrophages. Identification of the PCERA-1 receptor is expected to set up a new target for development of novel anti-inflammatory drugs. PUBLICATION ABSTRACT
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